Heterocyclic compounds and uses thereof

ABSTRACT

Compounds and pharmaceutical compositions that modulate kinase activity, including PI3 kinase activity, and compounds, pharmaceutical compositions, and methods of treatment of diseases and conditions associated with kinase activity, including PI3 kinase activity, are described herein.

CROSS REFERENCES TO RELATED APPLICATIONS

This application is a continuation application of U.S. application Ser.No. 14/661,656, filed Mar. 18, 2015, which claims priority to U.S.Provisional Application Nos. 61/955,717, filed Mar. 19, 2014,61/980,484, filed Apr. 16, 2014, 62/000,923, filed May 20, 2014,62/033,008, filed Aug. 4, 2014, 62/059,766, filed Oct. 3, 2014,62/075,173, filed Nov. 4, 2014, and 62/101,980, filed Jan. 9, 2015, theentireties of which are incorporated herein by reference.

BACKGROUND

The activity of cells can be regulated by external signals thatstimulate or inhibit intracellular events. The process by whichstimulatory or inhibitory signals are transmitted into and within a cellto elicit an intracellular response is referred to as signaltransduction. Over the past decades, cascades of signal transductionevents have been elucidated and found to play a central role in avariety of biological responses. Defects in various components of signaltransduction pathways have been found to account for a vast number ofdiseases, including numerous forms of cancer, inflammatory disorders,metabolic disorders, vascular and neuronal diseases (Gaestel et al.Current Medicinal Chemistry (2007) 14:2214-2234).

Kinases represent a class of important signaling molecules. Kinases cangenerally be classified into protein kinases and lipid kinases, andcertain kinases exhibit dual specificities. Protein kinases are enzymesthat phosphorylate other proteins and/or themselves (i.e.,autophosphorylation). Protein kinases can be generally classified intothree major groups based upon their substrate utilization: tyrosinekinases which predominantly phosphorylate substrates on tyrosineresidues (e.g., erb2, PDGF receptor, EGF receptor, VEGF receptor, src,abl), serine/threonine kinases which predominantly phosphorylatesubstrates on serine and/or threonine residues (e.g., mTorC1, mTorC2,ATM, ATR, DNA-PK, Akt), and dual-specificity kinases which phosphorylatesubstrates on tyrosine, serine and/or threonine residues.

Lipid kinases are enzymes that catalyze the phosphorylation of lipids.These enzymes, and the resulting phosphorylated lipids and lipid-derivedbiologically active organic molecules play a role in many differentphysiological processes, including cell proliferation, migration,adhesion, and differentiation. Certain lipid kinases are membraneassociated and they catalyze the phosphorylation of lipids contained inor associated with cell membranes. Examples of such enzymes includephosphoinositide(s) kinases (e.g., PI3-kinases, PI4-kinases),diacylglycerol kinases, and sphingosine kinases.

The phosphoinositide 3-kinases (PI3Ks) signaling pathway is one of themost highly mutated systems in human cancers. PI3K signaling is also akey factor in many other diseases in humans. PI3K signaling is involvedin many disease states including allergic contact dermatitis, rheumatoidarthritis, osteoarthritis, inflammatory bowel diseases, chronicobstructive pulmonary disorder, psoriasis, multiple sclerosis, asthma,disorders related to diabetic complications, and inflammatorycomplications of the cardiovascular system such as acute coronarysyndrome.

PI3Ks are members of a unique and conserved family of intracellularlipid kinases that phosphorylate the 3′-OH group onphosphatidylinositols or phosphoinositides. The PI3K family comprises 15kinases with distinct substrate specificities, expression patterns, andmodes of regulation. The class I PI3Ks (p110α, p110β, p110δ, and p110γ)are typically activated by tyrosine kinases or G-protein coupledreceptors to generate PIP3, which engages downstream effectors such asthose in the Akt/PDK1 pathway, mTOR, the Tec family kinases, and the Rhofamily GTPases. The class II and III PI3Ks play a key role inintracellular trafficking through the synthesis of PI(3)P and PI(3,4)P2.The PI3Ks are protein kinases that control cell growth (mTORC1) ormonitor genomic integrity (ATM, ATR, DNA-PK, and hSmg-1).

The delta (δ) isoform of class I PI3K has been implicated, inparticular, in a number of diseases and biological processes. PI3K-δ isexpressed primarily in hematopoietic cells including leukocytes such asT-cells, dendritic cells, neutrophils, mast cells, B-cells, andmacrophages. PI3K-δ is integrally involved in mammalian immune systemfunctions such as T-cell function, B-cell activation, mast cellactivation, dendritic cell function, and neutrophil activity. Due to itsintegral role in immune system function, PI3K-δ is also involved in anumber of diseases related to undesirable immune response such asallergic reactions, inflammatory diseases, inflammation mediatedangiogenesis, rheumatoid arthritis, and auto-immune diseases such aslupus, asthma, emphysema and other respiratory diseases. Other class IPI3K involved in immune system function includes PI3K-γ, which plays arole in leukocyte signaling and has been implicated in inflammation,rheumatoid arthritis, and autoimmune diseases such as lupus. Forexample, PI3K-γ and PI3K-δ are highly expressed in leukocytes and havebeen associated with adaptive and innate immunity; thus, these PI3Kisoforms can be important mediators in inflammatory disorders andhematologic malignancies.

The gamma (γ) isoform of class I PI3K consists of a catalytic subunitp110γ, which is associated with a p101 regulatory subunit. PI3K-γ isregulated by G protein-coupled receptors (GPCRs) via association withthe β/γ subunits of heterotrimeric G proteins. PI3K-γ is expressedprimarily in hematopoietic cells and cardiomyocytes and is involved ininflammation, the innate immune response, myeloid cell differentiation,immune cell trafficking, and mast cell function. Inhibitors of PI3K-γare useful for treating a variety of inflammatory diseases, allergies,and cardiovascular diseases, among others.

Unlike PI3K-δ, the beta (β) isoform of class I PI3K appears to beubiquitously expressed. PI3K-β has been implicated primarily in varioustypes of cancer including PTEN-negative cancer (Edgar et al. CancerResearch (2010) 70(3):1164-1172), and HER2-overexpressing cancer such asbreast cancer and ovarian cancer.

SUMMARY

Described herein are compounds capable of selectively inhibiting one ormore isoform(s) of class I PI3K without substantially affecting theactivity of the remaining isoforms of the same class. For example, insome embodiments, non-limiting examples of inhibitors capable ofselectively inhibiting PI3K-δ and/or PI3K-γ, but without substantiallyaffecting the activity of PI3K-α and/or PI3K-β are disclosed. In oneembodiment, the inhibitors provided herein can be effective inameliorating disease conditions associated with PI3K-δ and/or PI3K-γactivity. In one embodiment, the compounds are capable of selectivelyinhibiting PI3K-γ over PI3K-δ.

In one aspect, provided herein are compounds of Formula (I″) or (A″):

or an enantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof,wherein R^(3a), z, R^(1c), R^(2c), R¹, X, B, and W^(d) are definedherein.

In one aspect, provided herein are compounds of Formula (I′) or (A′):

or an enantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof,wherein R¹, X, B, and W^(d) are defined herein.

In one aspect, provided herein are compounds of Formula (I) or (A):

or an enantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof,wherein R¹, X, B, and W^(d) are defined herein.

In one embodiment, the compound of Formula (I″), (I′), (I), (A″), (A′),or (A) is predominately in an (S)-stereochemical configuration. In oneembodiment, the compound of Formula (I″), (I′), (I), (A″), (A′), or (A)is the S enantiomer having an enantiomeric excess selected from greaterthan about 25%, greater than about 55%, greater than about 80%, greaterthan about 90%, and greater than about 95%. In one embodiment, thecompound is present in a pharmaceutical composition comprising thecompound, or a pharmaceutically acceptable salt thereof, and one or morepharmaceutically acceptable excipients.

In certain embodiments, a compound disclosed herein selectivelymodulates PI3K gamma isoform. In certain embodiments, the compoundselectively inhibits the gamma isoform over the alpha or beta isoform.By way of non-limiting example, the ratio of selectivity can be greaterthan a factor of about 10, greater than a factor of about 50, greaterthan a factor of about 100, greater than a factor of about 200, greaterthan a factor of about 400, greater than a factor of about 600, greaterthan a factor of about 800, greater than a factor of about 1000, greaterthan a factor of about 1500, greater than a factor of about 2000,greater than a factor of about 5000, greater than a factor of about10,000, or greater than a factor of about 20,000, where selectivity canbe measured by ratio of IC₅₀ values, among other means. In oneembodiment, the selectivity of PI3K gamma isoform over PI3K alpha orbeta isoform is measured by the ratio of the IC₅₀ value against PI3Kalpha or beta isoform to the IC₅₀ value against PI3K gamma isoform.

In certain embodiments, a compound disclosed herein selectivelymodulates PI3K gamma isoform over the delta isoform. By way ofnon-limiting example, the ratio of selectivity can be greater than afactor of about 10, greater than a factor of about 50, greater than afactor of about 100, greater than a factor of about 200, greater than afactor of about 400, greater than a factor of about 600, greater than afactor of about 800, greater than a factor of about 1000, greater than afactor of about 1500, greater than a factor of about 2000, greater thana factor of about 5000, greater than a factor of about 10,000, orgreater than a factor of about 20,000, where selectivity can be measuredby ratio of IC₅₀ values, among other means. In one embodiment, theselectivity of PI3K gamma isoform over PI3K delta isoform is measured bythe ratio of the IC₅₀ value against PI3K delta isoform to the IC₅₀ valueagainst PI3K gamma isoform.

In certain embodiments, a compound as disclosed herein selectivelymodulates PI3K delta isoform. In certain embodiments, the compoundselectively inhibits the delta isoform over the alpha or beta isoform.By way of non-limiting example, the ratio of selectivity can be greaterthan a factor of about 10, greater than a factor of about 50, greaterthan a factor of about 100, greater than a factor of about 200, greaterthan a factor of about 400, greater than a factor of about 600, greaterthan a factor of about 800, greater than a factor of about 1000, greaterthan a factor of about 1500, greater than a factor of about 2000,greater than a factor of about 5000, greater than a factor of about10,000, or greater than a factor of about 20,000, where selectivity canbe measured by ratio of IC₅₀ values, among other means. In oneembodiment, the selectivity of PI3K delta isoform over PI3K alpha orbeta isoform is measured by the ratio of the IC₅₀ value against PI3Kalpha or beta isoform to the IC₅₀ value against PI3K delta isoform.

In certain embodiments, provided herein is a composition (e.g., apharmaceutical composition) comprising a compound described herein and apharmaceutically acceptable excipient. In some embodiments, providedherein is a method of inhibiting a PI3 kinase, comprising contacting thePI3 kinase with an effective amount of a compound or a pharmaceuticalcomposition described herein. In certain embodiments, a method isprovided for inhibiting a PI3 kinase wherein said PI3 kinase is presentin a cell. The inhibition can take place in a subject suffering from adisorder selected from cancer, bone disorder, inflammatory disease,immune disease, nervous system disease (e.g., a neuropsychiatricdisorder), metabolic disease, respiratory disease, thrombosis, andcardiac disease, among others. In certain embodiments, a secondtherapeutic agent is administered to the subject.

In certain embodiments, a method is provided for selectively inhibitinga PI3 kinase gamma isoform over PI3 kinase alpha or beta isoform whereinthe inhibition takes place in a cell. Non-limiting examples of themethods disclosed herein can comprise contacting PI3 kinase gammaisoform with an effective amount of a compound or a pharmaceuticalcomposition disclosed herein. In an embodiment, such contact can occurin a cell.

In certain embodiments, a method is provided for selectively inhibitinga PI3 kinase gamma isoform over PI3 kinase alpha or beta isoform whereinthe inhibition takes place in a subject suffering from a disorderselected from cancer, bone disorder, inflammatory disease, immunedisease, nervous system disease (e.g., a neuropsychiatric disorder),metabolic disease, respiratory disease, thrombosis, and cardiac disease,said method comprising administering an effective amount of a compoundor a pharmaceutical composition provided herein to said subject. Incertain embodiments, provided herein is a method of treating a subjectsuffering from a disorder associated with PI3 kinase, said methodcomprising selectively modulating the PI3 kinase gamma isoform over PI3kinase alpha or beta isoform by administering an amount of a compound ora pharmaceutical composition provided herein to said subject, whereinsaid amount is sufficient for selective modulation of PI3 kinase gammaisoform over PI3 kinase alpha or beta isoform.

In certain embodiments, a method is provided for selectively inhibitinga PI3 kinase delta isoform over PI3 kinase alpha or beta isoform whereinthe inhibition takes place in a cell. Non-limiting examples of themethods disclosed herein can comprise contacting PI3 kinase deltaisoform with an effective amount of a compound or a pharmaceuticalcomposition disclosed herein. In an embodiment, such contact can occurin a cell.

In certain embodiments, a method is provided for selectively inhibitinga PI3 kinase delta isoform over PI3 kinase alpha or beta isoform whereinthe inhibition takes place in a subject suffering from a disorderselected from cancer, bone disorder, inflammatory disease, immunedisease, nervous system disease (e.g., a neuropsychiatric disorder),metabolic disease, respiratory disease, thrombosis, and cardiac disease,said method comprising administering an effective amount of a compoundor a pharmaceutical composition provided herein to said subject. Incertain embodiments, provided herein is a method of treating a subjectsuffering from a disorder associated with PI3 kinase, said methodcomprising selectively modulating the PI3 kinase delta isoform over PI3kinase alpha or beta isoform by administering an amount of a compound ora pharmaceutical composition provided herein to said subject, whereinsaid amount is sufficient for selective modulation of PI3 kinase deltaisoform over PI3 kinase alpha or beta isoform.

In certain embodiments, a method is provided for selectively inhibitinga PI3 kinase gamma isoform over PI3 kinase delta isoform wherein theinhibition takes place in a cell. Non-limiting examples of the methodsdisclosed herein can comprise contacting PI3 kinase gamma isoform withan effective amount of a compound or a pharmaceutical compositiondisclosed herein. In an embodiment, such contact can occur in a cell.

In certain embodiments, a method is provided for selectively inhibitinga PI3 kinase gamma isoform over PI3 kinase delta isoform wherein theinhibition takes place in a subject suffering from a disorder selectedfrom cancer, bone disorder, inflammatory disease, immune disease,nervous system disease (e.g., a neuropsychiatric disorder), metabolicdisease, respiratory disease, thrombosis, and cardiac disease, saidmethod comprising administering an effective amount of a compound or apharmaceutical composition provided herein to said subject. In certainembodiments, provided herein is a method of treating a subject sufferingfrom a disorder associated with PI3 kinase, said method comprisingselectively modulating the PI3 kinase gamma isoform over PI3 kinasedelta isoform by administering an amount of a compound or apharmaceutical composition provided herein to said subject, wherein saidamount is sufficient for selective modulation of PI3 kinase gammaisoform over PI3 kinase delta isoform.

In certain embodiments, provided herein is a method of inhibiting a PI3kinase in a subject suffering from an inflammatory disease, an immunedisease, or a respiratory disease, comprising administering to thesubject an effective amount of a compound provided herein (e.g., acompound of any of Formulae (I″), (I′), (A′), (I), (A), (II), (III),(IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII), (XIV), (XV),(XVI), and (XVII), e.g., as described herein). In one embodiment, thecompound is a selective PI3K-γ inhibitor, e.g., Compound 4. In oneembodiment, the subject is a mammal. In one embodiment, the mammal is ahuman. In one embodiment, the subject is a human.

In some embodiments, the disorder treated by the methods or compoundsdisclosed herein is a cancer. In some embodiments, the cancer is a solidor soft tissue tumor (e.g., a carcinoid, carcinoma or sarcoma), ahematopoietic tissue tumor (e.g., a heme malignancy), or a metastaticlesion, e.g., a metastatic lesion of any of the cancers or tumorsdisclosed herein. In one embodiment, the cancer is metastatic cancer tothe bone.

In one embodiment, the cancer treated by the methods or compoundsdisclosed herein is a a soft tissue tumor, a heme malignancy, or ahematological cancer. In one embodiment, the cancer is acute myeloidleukemia (AML), chronic myeloid leukemia (CML), myelodysplastic syndrome(MDS), myeloproliferative disorders, mast cell cancer, Hodgkin disease,non-Hodgkin lymphomas, diffuse large B-cell lymphoma, humanlymphotrophic virus type 1 (HTLV-1) leukemia/lymphoma, AIDS-relatedlymphoma, adult T-cell lymphoma, acute lymphoblastic leukemia (ALL),T-cell acute lymphoblastic leukemia, B-cell acute lymphoblasticleukemia, chronic lymphocytic leukemia, or multiple myeloma (MM). In oneembodiment, the cancer is leukemia or lymphoma. In one embodiment, theleukemia is B-cell acute lymphoblastic leukemia (B-ALL), acute myeloidleukemia (AML), acute lymphoblastic leukemia, chronic myeloid leukemia,hairy cell leukemia, myeloproliferative disorders, acute myelogenousleukemia (AML), chronic myelogenous leukemia (CML), chronic lymphocyticleukemia (CLL), multiple myeloma (MM), myelodysplastic syndrome (MDS),or mast cell cancer. In one embodiment, the lymphoma is diffuse largeB-cell lymphoma, B-cell immunoblastic lymphoma, small non-cleaved cellor Burkitt lymphoma, human lymphotropic virus-type 1 (HTLV-1)leukemia/lymphoma, adult T-cell lymphoma, Hodgkin disease, ornon-Hodgkin lymphomas, or a metastatic lesion thereof.

In one embodiment, the cancer treated by the methods or compoundsdisclosed herein is a solid tumor (e.g., a carcinoid, carcinoma orsarcoma), or a metastatic lesion thereof. In one embodiment, the canceris a lung cancer (e.g., non-small cell lung cancer or small cell lungcancer); a skin cancer; a melanoma; a prostate cancer; a glioblastoma;an endometrial cancer; a pancreatic cancer (e.g., pancreaticadenocarcinoma (e.g., pancreatic ductal adenocarcinoma (PDA)); a renalcell carcinoma; a colorectal cancer; a breast cancer (e.g., triplenegative breast cancer); a thyroid cancer; a sarcoma, a liver orhepatocellular cancer (HCC), a head and neck cancer, a cervical orvulvar cancer, an esophageal cancer, a gastric cancer, an adrenalcancer, or an ovarian cancer, or a metastatic lesion thereof. In oneembodiment, the solid tumor is prostate cancer, breast cancer, or aglioblastoma, or a metastatic lesion thereof.

In some embodiments, the cancer or tumor treated is a solid, fibrotictumor chosen from one or more of pancreatic (e.g., pancreaticadenocarcinoma or pancreatic ductal adenocarcinoma), breast, colorectal,colon, lung (e.g., a small or non-small cell lung cancer), skin,ovarian, prostate, cervix, gastrointestinal (e.g., carcinoid orstromal), stomach, head and neck, kidney, brain cancer, or a metastaticlesion thereof.

In some embodiments, the cancer or tumor treated using the methods orcompounds disclosed herein is a cancer or tumor chosen from one or moreof the head, neck, nasal cavity, paranasal sinuses, nasopharynx, oralcavity, oropharynx, larynx, hypopharynx, salivary glands,paragangliomas, pancreas, stomach, skin, esophagus, endometrium, liverand biliary tree, bone, intestine, colon, rectum, ovaries, prostate,lung, breast, lymphatic system, blood, bone marrow central nervoussystem, brain, or a metastatic lesion thereof.

In some embodiments, the disorder treated by the methods or compoundsdisclosed herein is an inflammatory disease or an immune disease. In oneembodiment, the inflammatory disease or the immune disease is asthma,emphysema, allergy, dermatitis, arthritis (e.g., rheumatoid arthritis),psoriasis, lupus erythematosus, graft versus host disease, inflammatorybowel disease, eczema, scleroderma, Crohn's disease, or multiplesclerosis. In one embodiment, the disorder is rheumatoid arthritis. Inone embodiment, the disorder is rheumatoid arthritis, and the amount ofthe compound is effective to ameliorate one or more symptoms associatedwith rheumatoid arthritis, wherein the symptom associated withrheumatoid arthritis is independently a reduction in the swelling of thejoints, a reduction in serum anti collagen levels, a reduction in boneresorption, a reduction in cartilage damage, a reduction in pannus, or areduction in inflammation.

In some embodiments, the disorder treated by the methods or compoundsdisclosed herein is a respiratory disease. In one embodiment, therespiratory disease is asthma, chronic obstructive pulmonary disease(COPD), chronic bronchitis, emphysema, or bronchiectasis. In oneembodiment, the disorder is asthma.

In certain embodiments, a method is provided for selectively inhibitinga PI3 kinase gamma isoform over PI3 kinase alpha or beta isoform whereinthe inhibition takes place in a subject suffering from a respiratorydisease. In one embodiment, the respiratory disease is asthma, chronicobstructive pulmonary disease (COPD), chronic bronchitis, emphysema, orbronchiectasis. In one embodiment, the respiratory disease is asthma. Inone embodiment, the respiratory disease is COPD. In one embodiment, themethod further comprises administration of one or more therapeuticagents selected from chemotherapeutic agents, cytotoxic agents, andradiation. In one embodiment, the compound is administered incombination with an mTOR inhibitor. In one embodiment, the compound isadministered in combination with one or more of: an agent that inhibitsIgE production or activity,2-(4-(6-cyclohexyloxy-2-naphtyloxy)phenylacetamide)benzoic acid, an mTORinhibitor, rapamycin, a TORC1 inhibitor, a TORC2 inhibitor, an anti-IgEantibody, prednisone, corticosteroid, a leukotriene inhibitor, XOLAIR,ADVAIR, SINGULAIR, or SPIRIVA. In one embodiment, the compound isadministered in combination with one or more of: a mitotic inhibitor, analkylating agent, an anti-metabolite, an intercalating antibiotic, agrowth factor inhibitor, a cell cycle inhibitor, an enzyme, atopoisomerase inhibitor, an anti-hormone, an angiogenesis inhibitor, ananti-androgen, or an anti-receptor kinase antibody. In one embodiment,the compound is administered in combination with one or more of:Imatinib Mesylate, bortezomib, bicalutamide, gefitinib, ADRIAMYCIN,alkylating agents, alkyl sulfonates, ethylenimines, altretamine,triethylenemelamine, trietylenephosphoramide,triethylenethiophosphaoramide, trimethylolomelamine, nitrogen mustards,chlorambucil, chlornaphazine, cholophosphamide, estramustine,ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride,melphalan, novembichin, phenesterine, prednimustine, trofosfamide,uracil mustard, nitrosureas, antibiotics, anti-metabolites, denopterin,methotrexate, pteropterin, trimetrexate, 5-fluorouracil (5-FU),fludarabine, 6-mercaptopurine, thiamiprine, thioguanine, ancitabine,azacitidine, 6-azauridine, carmofur, cytarabine, dideoxyuridine,doxifluridine, enocitabine, floxuridine, androgens, anti-adrenals, folicacid replenisher, arabinoside, cyclophosphamide, thiotepa, taxanes,anti-hormonal agents, anti-estrogens, tamoxifen, raloxifene, aromataseinhibiting 4(5)-imidazoles, 4-hydroxytamoxifen, trioxifene, keoxifene,onapristone, toremifene, anti-androgens, chlorambucil, gemcitabine,6-thioguanine; mercaptopurine; cisplatin, carboplatin, vincristine;vinorelbine, vinblastin, ifosfamide, mitomycin C, daunorubicin,doxorubicin, mitoxantrone, HERCEPTIN, AVASTIN, ERBITUX, RITUXAN, TAXOL,ARIMIDEX, TAXOTERE, or an anti-receptor tyrosine kinase antibodyselected from cetuximab, panitumumab, trastuzumab, anti CD20 antibody,rituximab, tositumomab, alemtuzumab, bevacizumab, obinutuzumab (GAZYVA),and gemtuzumab. In one embodiment, the compound is administered incombination with one or more of: bortezomib, ADRIAMYCIN, alkylatingagents, anti-metabolites, denopterin, pteropterin, trimetrexate, anitrogen mustard, chlorambucil, chlornaphazine, cholophosphamide,estramustine, ifosfamide, mechlorethamine, mechlorethamine oxidehydrochloride, melphalan, novembichin, phenesterine, prednimustine,trofosfamide, uracil mustard, methotrexate, fludarabine,6-mercaptopurine, thiamiprine, thioguanine, ancitabine, azacitidine,6-azauridine, carmofur, cytarabine, dideoxyuridine, doxifluridine,enocitabine, floxuridine, androgens, cyclophosphamide, taxanes,anti-hormonal agents, gemcitabine; cisplatin, carboplatin, vincristine,vinorelbine, vinblastin, ifosfamide, mitomycin C, daunorubicin,doxorubicin, mitoxantrone, HERCEPTIN, AVASTIN, ERBITUX, RITUXAN, TAXOL,ARIMIDEX, or TAXOTERE. In one embodiment, the compound is administeredin combination with one or more of: non-steroidal anti-inflammatorydrugs (NSAIDs), corticosteroids, prednisone, chloroquine,hydroxychloroquine, azathioprine, cyclophosphamide, methotrexate,cyclosporine, anti-CD20 antibodies, ENBREL, REMICADE, HUMIRA, AVONEX, orREBIF.

In one embodiment, provided herein is a method of inhibiting a PI3kinase in a subject suffering from a cancer, comprising administering tothe subject an effective amount of a compound provided herein (e.g., acompound of any of Formulae (I″), (I′), (A′), (I), (A), (II), (III),(IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII), (XIV), (XV),(XVI), and (XVII), e.g., as described herein). In one embodiment, thecompound is a selective PI3K-γ inhibitor, e.g., Compound 4. In oneembodiment, the cancer is selected from acute myeloid leukemia (AML),chronic myeloid leukemia (CML), myelodysplastic syndrome (MDS),myeloproliferative disorders, mast cell cancer, Hodgkin disease,non-Hodgkin lymphomas, diffuse large B-cell lymphoma, humanlymphotrophic virus-type 1 (HTLV-1) leukemia/lymphoma, AIDS-relatedlymphoma, adult T-cell lymphoma, acute lymphoblastic leukemia (ALL),B-cell acute lymphoblastic leukemia, T-cell acute lymphoblasticleukemia, chronic lymphocytic leukemia, or multiple myeloma (MM). In oneembodiment, the cancer is leukemia or lymphoma. In one embodiment, theleukemia is selected from B-cell acute lymphoblastic leukemia (B-ALL),acute lymphocytic leukemia, hairy cell leukemia, myelodysplasia,myeloproliferative disorders, acute myelogenous leukemia (AML), chronicmyelogenous leukemia (CML), chronic lymphocytic leukemia (CLL), multiplemyeloma (MM), myelodysplastic syndrome (MDS), or mast cell cancer. Inone embodiment, the lymphoma is selected from diffuse large B-celllymphoma, B-cell immunoblastic lymphoma, small non-cleaved cell orBurkitt lymphoma, human lymphotropic virus-type 1 (HTLV-1)leukemia/lymphoma, AIDS-related lymphoma, adult T-cell lymphoma, Hodgkindisease, or non-Hodgkin lymphomas. In one embodiment, the compound isadministered in combination with one or more therapeutic agents providedherein.

In one embodiment, provided herein is a method of inhibiting a PI3kinase in a subject suffering from an inflammatory disease or an immunedisease, comprising administering to the subject an effective amount ofa compound provided herein (e.g., a compound of any of Formulae (I″),(I′), (A′), (I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X),(XI), (XII), (XIII), (XIV), (XV), (XVI), and (XVII), e.g., as describedherein). In one embodiment, the compound is a selective PI3K-γinhibitor, e.g., Compound 4. In one embodiment, the inflammatory diseaseor immune disease is asthma, emphysema, allergy, dermatitis, rheumatoidarthritis, psoriasis, lupus erythematosus, graft versus host disease,inflammatory bowel disease, eczema, scleroderma, Crohn's disease, ormultiple sclerosis. In one embodiment, the inflammatory disease orimmune disease is rheumatoid arthritis. In one embodiment, the compoundis administered in combination with one or more therapeutic agentsprovided herein.

In one embodiment, provided herein is a method of inhibiting a PI3kinase in a subject suffering from a respiratory disease, comprisingadministering to the subject an effective amount of a compound providedherein (e.g., a compound of Formula I). In one embodiment, therespiratory disease is asthma, chronic obstructive pulmonary disease(COPD), chronic bronchitis, emphysema, or bronchiectasis. In oneembodiment, the respiratory disease is asthma. In one embodiment, thecompound is administered in combination with one or more therapeuticagents provided herein.

In certain embodiments, provided herein is a method of inhibiting PI3K-γin a subject, comprising administering to the subject an effectiveamount of a compound provided herein (e.g., a compound of any ofFormulae (I″), (I′), (A′), (I), (A), (II), (III), (IV), (V), (VI),(VII), (IX), (X), (XI), (XII), (XIII), (XIV), (XV), (XVI), and (XVII),e.g., as described herein). In one embodiment, the compound is aselective PI3K-γ inhibitor, e.g., Compound 4.

In certain embodiments, provided herein is a method of making a compounddescribed herein.

In certain embodiments, provided herein is a reaction mixture comprisinga compound described herein.

In certain embodiments, provided herein is a kit comprising a compounddescribed herein.

In some embodiments, a method is provided for treating a disease ordisorder described herein, the method comprising administering atherapeutically effective amount of a compound or a pharmaceuticalcomposition described herein to a subject.

In some embodiments, a method is provided for treating a PI3K mediateddisorder in a subject, the method comprising administering atherapeutically effective amount of a compound or a pharmaceuticalcomposition described herein to a subject.

In some embodiments, provided herein is a use of a compound or apharmaceutical composition described herein for the treatment of adisease or disorder described herein in a subject.

In some embodiments, provided herein is a use of a compound or apharmaceutical composition described herein for the treatment of a PI3Kmediated disorder in a subject.

In some embodiments, provided herein is a use of a compound or apharmaceutical composition described herein in the manufacture of amedicament for the treatment of a disease or disorder described hereinin a subject.

In some embodiments, provided herein is use of a compound or apharmaceutical composition described herein in the manufacture of amedicament for the treatment of a PI3K mediated disorder in a subject.

Additional features or embodiments of the compounds, compositions ormethods described herein include one or more of the following:

In some embodiments, the compound is a PI3K-gamma inhibitor, e.g., aselective PI3K-gamma inhibitor, or a pharmaceutically acceptable formthereof.

In some embodiments, the compound is a compound of any of Formulae (I″),(I′), (A′), (I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X),(XI), (XII), (XIII), (XIV), (XV), (XVI), and (XVII), or apharmaceutically acceptable form thereof. In one embodiment, thecompound is a compound of Formula (XVII). In one embodiment, the one ormore compounds are PI3K gamma inhibitors, e.g., selective PI3K-gammainhibitors.

In some embodiments, the compound is chosen from one or more ofcompounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41,52, 60, 61, 63, 73, 75, 77, 79, 80, 81, or 88, or a pharmaceuticallyacceptable form thereof. In one embodiment, the one or more compoundsare PI3K gamma inhibitors, e.g., selective PI3K-gamma inhibitors.

In one embodiment, the compound is a compound of Formula (I″), (I′), or(I), or a pharmaceutically acceptable form thereof. In one embodiment,the compound is a compound of Formula I, or a pharmaceuticallyacceptable form thereof.

In one embodiment, the compound is Compound 4 (also referred to hereinas “Compound BB”):

or a pharmaceutically acceptable form thereof. In some embodiments,Compound 4 is a PI3K gamma inhibitor, e.g., a selective PI3K-gammainhibitor.

In some embodiments of the methods or uses disclosed herein, the PI3Kgamma inhibitor, e.g., the selective PI3K-gamma inhibitor, or a compoundas described herein (e.g., compound of any of Formulae (I″), (I′), (A′),(I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI), (XII),(XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound 4), isadministered to a subject at a dose (e.g., a therapeutically effectivedose) of about 2 mg, 1-3 mg, 1-5 mg, 1-10 mg, 0.5-20 mg, or 0.1-50 mg.In some embodiments, the dose (e.g., a therapeutically effective dose)is about 2 mg, 1-3 mg, 1-5 mg, 1-10 mg, 0.5-20 mg, 0.1-50 mg, 0.1-75 mg,0.5-75 mg, 1-75 mg, 0.1-100 mg, 0.5-100 mg, or 1-100 mg. In someembodiments, the dose is about 1-10 mg. In some embodiments, the dose isabout 1-50 mg. In some embodiments, the dose is about 1-100 mg.

In some embodiments of the methods or uses disclosed herein, the PI3Kgamma inhibitor, e.g., the selective PI3K-gamma inhibitor, or a compoundas described herein (e.g., a compound of any of Formulae (I″), (I′),(A′), (I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI),(XII), (XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound 4), isadministered to a subject at a dose (e.g., a therapeutically effectivedose) of about 0.029 mg/kg, 0.014-0.14 mg/kg, 0.02-0.04 mg/kg, 0.01-0.05mg/kg, 0.01-0.1, or 0.01-0.5 mg/kg. In one embodiment, theadministration is intra-tracheally.

In some embodiments of the methods or uses disclosed herein, the PI3Kgamma inhibitor, e.g., the selective PI3K-gamma inhibitor, or a compoundas described herein (e.g., a compound of any of Formulae (I″), (I′),(A′), (I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI),(XII), (XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound 4), isadministered to a subject at a treatment schedule chosen from, e.g.,once every two days, once per day, or twice per day.

In some embodiments of the methods or uses disclosed herein, the PI3Kgamma inhibitor, e.g., the selective PI3K-gamma inhibitor, or a compoundas described herein (e.g., a compound of any of Formulae (I″), (I′),(A′), (I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI),(XII), (XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound 4) isadministered at a dose such that it selectively inhibits PI3K-gamma butachieves less than 10% or 20% inhibition of PI3K-delta.

In some embodiments of the methods or uses disclosed herein, the PI3Kgamma inhibitor, e.g., the selective PI3K-gamma inhibitor, or a compoundas described herein (e.g., a compound of any of Formulae (I″), (I′),(A′), (I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI),(XII), (XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound 4) isadministered at a dose such that the compound's level in the subject'sblood does not rise higher than a predetermined level, e.g., the IC50 ofPI3K-delta, within a selected time period, e.g., 24 hours. In someembodiments of the methods or uses disclosed herein, the PI3K gammainhibitor or compound is administered at a dose such that the compound'slevel in the subject's blood does not rise higher than a predeterminedlevel, e.g., the IC20 of PI3K-delta, within a selected time period,e.g., 24 hours.

In some embodiments of the methods or uses disclosed herein, the PI3Kgamma inhibitor, e.g., the selective PI3K-gamma inhibitor, or a compoundas described herein (e.g., a compound of any of Formulae (I″), (I′),(A′), (I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI),(XII), (XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound 4), isadministered to a subject in an amount such that the level of thecompound in the subject's body is above the IC50 of PI3K-gamma during atleast 70%, 80%, 90%, 95%, 97%, 98%, 99%, or 99% of selected time period,e.g., 24 hours, immediately following the administration.

In some embodiments of the methods or uses disclosed herein, the PI3Kgamma inhibitor, e.g., the selective PI3K-gamma inhibitor, or a compoundas described herein (e.g., a compound of any of Formulae (I″), (I′),(A′), (I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI),(XII), (XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound 4), isadministered to a subject in an amount such that the level of thecompound in the subject's body is above the IC90 of PI3K-gamma during atleast 50%, 60%, 70%, 80%, 90%, 95%, 97%, 98%, 99%, or 99% of a selectedtime period, e.g., 24 hours, immediately following the administration.

In some embodiments of the methods or uses disclosed herein, the subjectis a human and the PI3K gamma inhibitor, e.g., the selective PI3K-gammainhibitor, or a compound as described herein (e.g., a compound of any ofFormulae (I″), (I′), (A′), (I), (A), (II), (III), (IV), (V), (VI),(VII), (IX), (X), (XI), (XII), (XIII), (XIV), (XV), (XVI), and (XVII),e.g., Compound 4) has a half life of about 8-15 hours, or about 10-13hours.

In some embodiments of the methods or uses disclosed herein, treatmentwith a PI3K gamma inhibitor, e.g., the selective PI3K-gamma inhibitor,or a compound as described herein (e.g., a compound of any of Formulae(I″), (I′), (A′), (I), (A), (II), (III), (IV), (V), (VI), (VII), (IX),(X), (XI), (XII), (XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound4) is more effective than treatment with a PI3K delta selectiveinhibitor, e.g., when both inhibitors are administered at the same doseand/or through the same route of administration.

In some embodiments of the methods or uses disclosed herein, treatmentwith a PI3K gamma inhibitor, e.g., the selective PI3K-gamma inhibitor,or a compound as described herein (e.g., a compound of any of Formulae(I″), (I′), (A′), (I), (A), (II), (III), (IV), (V), (VI), (VII), (IX),(X), (XI), (XII), (XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound4) is more effective (e.g., at least 10%, 20%, 40%, 60%, or 80% moreeffective) than treatment with a PI3K delta selective inhibitor, e.g.,when both inhibitors are administered at the same dose and/or throughthe same route of administration.

In some embodiments of the methods or uses disclosed herein, the subjectis refractory or has failed to respond to a PI3K-δ inhibitor.

Inflammatory Diseases

Methods of treating or preventing an inflammatory disease in a subjectusing the PI3K gamma inhibitor, e.g., the selective PI3K-gammainhibitor, or a compound as described herein (e.g., a compound of any ofFormulae (I″), (I′), (A′), (I), (A), (II), (III), (IV), (V), (VI),(VII), (IX), (X), (XI), (XII), (XIII), (XIV), (XV), (XVI), and (XVII),e.g., Compound 4, as described herein) are disclosed. In certainembodiments, the inflammatory disease is chosen from COPD, arthritis,asthma, psoriasis, scleroderma, myositis, sarcoidosis, dermatomyositis,CREST syndrome, systemic lupus erythematosus, Sjorgren syndrome,encephalomyelitis, or inflammatory bowel disease (IBD). In someembodiments, inflammatory disease is COPD or arthritis.

In certain embodiments, the PI3K gamma inhibitor, e.g., the selectivePI3K-gamma inhibitor, or a compound as described herein (e.g., acompound of any of Formulae (I″), (I′), (A′), (I), (A), (II), (III),(IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII), (XIV), (XV),(XVI), and (XVII), e.g., Compound 4, as described herein) isadministered at a dose such that it selectively inhibits PI3K-gamma butachieves less than 10% or 20% inhibition of PI3K-delta.

In certain embodiments, the PI3K gamma inhibitor, e.g., the selectivePI3K-gamma inhibitor, or a compound as described herein (e.g., acompound of any of Formulae (I″), (I′), (A′), (I), (A), (II), (III),(IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII), (XIV), (XV),(XVI), and (XVII), e.g., Compound 4, as described herein) isadministered at a dose such that the compound's level in the subject'sblood does not rise higher than a predetermined level, e.g., the IC50 ofPI3K-delta, within a selected time period, e.g., 24 hours. In certainembodiments, the PI3K gamma inhibitor or a compound, e.g., Compound 4,as described herein) is administered at a dose such that the compound'slevel in the subject's blood does not rise higher than a predeterminedlevel, e.g., the IC20 of PI3K-delta, within a selected time period,e.g., 24 hours.

In some embodiments of the methods or uses disclosed herein, the subjectis refractory or has failed to respond to a PI3K-δ inhibitor.

In certain embodiments, a method of treating or preventing arthritis, orperiosteal bone formation, in a subject is disclosed. The methodincludes administering to the subject a PI3K gamma inhibitor, e.g., theselective PI3K-gamma inhibitor, or a compound as described herein (e.g.,a compound of any of Formulae (I″), (I′), (A′), (I), (A), (II), (III),(IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII), (XIV), (XV),(XVI), and (XVII), e.g., Compound 4, as described herein), in atherapeutically effective amount, e.g., an amount sufficient to treat orprevent the arthritis. In certain embodiments, the subject has or is atrisk of having, or is identified as having or being at risk of having,periosteal bone formation.

In certain embodiments, the subject has or is identified as havingreduced periosteal bone formation after administration of the PI3K gammainhibitor or a compound as described herein, e.g., the subject shows areduction in periosteal bone formation or periosteal bone width that isreduced by at least 10%, 20%, 40%, 47%, 50%, 52%, 60%, 80%, or 82%compared to a reference value (e.g., an untreated control), afteradministration of the PI3K gamma inhibitor or a compound as describedherein.

In certain embodiments, the therapeutically effective amount is anamount sufficient to reduce a periosteal bone formation or periostealbone width by at least 10%, 20%, 40%, 47%, 50%, 52%, 60%, 80%, or 82%compared to a reference value, after administration of the PI3K gammainhibitor or a compound as described herein.

In certain embodiments, the subject has, or is identified as having, ahistopathology score for periosteal bone formation or periosteal bonewidth that is reduced by at least 1, 2, or 3 points compared to areference value, after administration of the PI3K gamma inhibitor or acompound as described herein.

In certain embodiments, the therapeutically effective amount is anamount sufficient to reduce a histopathology score for periosteal boneformation or periosteal bone width by at least 1, 2, or 3 pointscompared to a reference value, after administration of the PI3K gammainhibitor or a compound as described herein.

In certain embodiments, the subject has, or is identified as having, aninflammation that is reduced by at least 10%, 20%, 27%, 30%, 36%, 40%,45%, 50%, or 57% compared to a reference value, after administration ofthe PI3K gamma inhibitor or a compound as described herein.

In certain embodiments, the therapeutically effective amount is anamount that is sufficient to reduce an inflammation by at least 10%,20%, 27%, 30%, 36%, 40%, 45%, 50%, or 57% compared to a reference value,after administration of the PI3K gamma inhibitor or a compound asdescribed herein.

In certain embodiments, the subject has, or is identified as having, ahistopathology score for inflammation that is reduced by at least 1, 2,or 3 points compared to a reference value, after administration of thePI3K gamma inhibitor or a compound as described herein.

In certain embodiments, the therapeutically effective amount is anamount that is sufficient to reduce a histopathology score forinflammation by at least 1, 2, or 3 points compared to a referencevalue, after administration of the PI3K gamma inhibitor or a compound asdescribed herein.

In certain embodiments, the subject has, or is identified as having,pannus that is reduced by at least 10%, 20%, 28%, 30%, 40%, 44%, 50%, or60%, 70%, or 71% compared to a reference value, after administration ofthe PI3K gamma inhibitor or a compound as described herein.

In certain embodiments, the therapeutically effective amount is anamount that is sufficient to reduce pannus by at least 10%, 20%, 28%,30%, 40%, 44%, 50%, or 60%, 70%, or 71% compared to a reference value,after administration of the PI3K gamma inhibitor or a compound asdescribed herein.

In certain embodiments, the subject has, or is identified as having, ahistopathology score for pannus that is reduced by at least 1, 2, or 3points compared to a reference value, after administration of the PI3Kgamma inhibitor or a compound as described herein.

In certain embodiments, the therapeutically effective amount is anamount that is sufficient to reduce a histopathology score for pannus byat least 1, 2, or 3 points compared to a reference value, afteradministration of the PI3K gamma inhibitor or a compound as describedherein.

In certain embodiments, the subject has, or is identified as having,cartilage damage that is reduced by at least 10%, 20%, 28%, 30%, 40%,45%, 50%, or 59% compared to a reference value, after administration ofthe PI3K gamma inhibitor or a compound as described herein.

In certain embodiments, the therapeutically effective amount is anamount that is sufficient to reduce cartilage damage by at least 10%,20%, 28%, 30%, 40%, 45%, 50%, or 59% compared to a reference value,after administration of the PI3K gamma inhibitor or a compound asdescribed herein.

In certain embodiments, the subject has, or is identified as having, ahistopathology score for cartilage damage that is reduced by at least 1,2, or 3 points compared to a reference value, after administration ofthe PI3K gamma inhibitor or a compound as described herein.

In certain embodiments, the therapeutically effective amount is anamount that is sufficient to reduce a histopathology score for cartilagedamage by at least 1, 2, or 3 points compared to a reference value,after administration of the PI3K gamma inhibitor or a compound asdescribed herein.

In certain embodiments, the subject has, or is identified as having,bone resorption that is reduced by at least 10%, 20%, 25%, 30%, 40%,44%, 50%, 60%, or 65% compared to a reference value, afteradministration of the PI3K gamma inhibitor or a compound as describedherein.

In certain embodiments, the therapeutically effective amount is anamount that is sufficient to reduce bone resorption by at least 10%,20%, 25%, 30%, 40%, 44%, 50%, 60%, or 65% compared to a reference value,after administration of the PI3K gamma inhibitor.

In certain embodiments, the subject has, or is identified as having, ahistopathology score for bone resorption that is reduced by at least 1,2, or 3 points compared to a reference value, after administration ofthe PI3K gamma inhibitor or a compound as described herein.

In certain embodiments, the therapeutically effective amount is anamount that is sufficient to reduce a histopathology score for boneresorption by at least 1, 2, or 3 points compared to a reference value,after administration of the PI3K gamma inhibitor or a compound asdescribed herein.

In certain embodiments, the subject has, or is identified as having,joint swelling that is reduced compared to a reference value, afteradministration of the PI3K gamma inhibitor or a compound as describedherein.

In certain embodiments, the therapeutically effective amount is anamount that is sufficient to reduce joint swelling compared to areference value, after administration of the PI3K gamma inhibitor or acompound as described herein.

In certain embodiments, the subject has, or is identified as having, ahistopathology score for joint swelling that is reduced compared to areference value, after administration of the PI3K gamma inhibitor or acompound as described herein.

In certain embodiments, the subject has, or is identified as having, ahistopathology score for joint swelling that is reduced compared to areference value, after administration of the PI3K gamma inhibitor or acompound as described herein.

In certain embodiments, the subject has, or is identified as having, ananti-collagen level that is reduced compared to a reference value, afteradministration of the PI3K gamma inhibitor.

In certain embodiments, the therapeutically effective amount is anamount that is sufficient to reduce an anti-collagen level compared to areference value, after administration of the PI3K gamma inhibitor or acompound as described herein.

In certain embodiments, the subject has, or is identified as having, ahistopathology score for an anti-collagen level that is reduced comparedto a reference value, after administration of the PI3K gamma inhibitor.

In certain embodiments, the therapeutically effective amount is anamount sufficient to reduce an anti-collagen level compared to areference value, after administration of the PI3K gamma inhibitor or acompound as described herein.

In certain embodiments, the PI3K gamma inhibitor or a compound asdescribed herein (e.g., a compound of any of Formulae (I″), (I′), (A′),(I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI), (XII),(XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound 4) isadministered at a dose such that it selectively inhibits PI3K-gamma butachieves less than 10% or 20% inhibition of PI3K-delta.

In certain embodiments, the PI3K gamma inhibitor or a compound asdescribed herein (e.g., a compound of any of Formulae (I″), (I′), (A′),(I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI), (XII),(XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound 4) isadministered at a dose such that the compound's level in the subject'sblood does not rise higher than a predetermined level, e.g., the IC50 ofPI3K-delta, within a selected time period, e.g., 24 hours. In certainembodiments, the PI3K gamma inhibitor or a compound as described herein(e.g., a compound of any of Formulae (I″), (I′), (A′), (I), (A), (II),(III), (IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII), (XIV),(XV), (XVI), and (XVII), e.g., Compound 4) is administered at a dosesuch that the compound's level in the subject's blood does not risehigher than a predetermined level, e.g., the IC20 of PI3K-delta, withina selected time period, e.g., 24 hours.

In other embodiments, a method of reducing neutrophil migration orinfiltration in a subject suffering from an inflammatory disease isprovided. The method includes administering to the subject a PI3K gammainhibitor or a compound as described herein (e.g., a compound of any ofFormulae (I″), (I′), (A′), (I), (A), (II), (III), (IV), (V), (VI),(VII), (IX), (X), (XI), (XII), (XIII), (XIV), (XV), (XVI), and (XVII),e.g., Compound 4, as described herein), in an amount sufficient toreduce or inhibit the neutrophil migration or infiltration in thesubject.

In certain embodiments, the subject has, or is identified as having,neutrophil migration that is reduced by at least about 10%, 20%, 40%,60%, 80%, or 90% compared to a reference value, after administration ofthe PI3K gamma inhibitor or a compound as described herein.

In certain embodiments, the therapeutically effective amount is anamount that is sufficient to reduce neutrophil migration by at leastabout 10%, 20%, 40%, 60%, 80%, or 90% compared to a reference value,after administration of the PI3K gamma inhibitor or a compound asdescribed herein.

Cancer

Methods of treating or preventing a cancer in a subject using the PI3Kgamma inhibitor or a compound as described herein (e.g., a compound ofany of Formulae (I″), (I′), (A′), (I), (A), (II), (III), (IV), (V),(VI), (VII), (IX), (X), (XI), (XII), (XIII), (XIV), (XV), (XVI), and(XVII), e.g., Compound 4, as described herein) are disclosed. In certainembodiments, the cancer is, or is identified as being, a solid tumor(e.g., lung cancer, melanoma, breast cancer, sarcoma, hepatocellularcancer, head and neck cancer, cervical or vulvar cancer, esophagealcancer, gastric cancer, adrenal cancer, colon cancer, or glioblastoma)or a hematologic cancer (e.g., a chronic lymphocytic leukemia (CLL)),e.g., as described herein. In one embodiment, the cancer is melanoma,bladder cancer, head and neck cancer, lung cancer (e.g., non-small celllung cancer), renal cell carcinoma, ovarian cancer, breast cancer (e.g.,triple-negative breast cancer), colon cancer, or glioblastoma.

In certain embodiments, the PI3K gamma inhibitor or a compound asdescribed herein (e.g., a compound of any of Formulae (I″), (I′), (A′),(I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI), (XII),(XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound 4, as describedherein) is administered at a dose such that it selectively inhibitsPI3K-gamma but achieves less than 10% or 20% inhibition of PI3K-delta.

In certain embodiments, the PI3K gamma inhibitor or a compound asdescribed herein (e.g., a compound of any of Formulae (I″), (I′), (A′),(I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI), (XII),(XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound 4, as describedherein) is administered at a dose such that the compound's level in thesubject's blood does not rise higher than a predetermined level, e.g.,the IC50 of PI3K-delta, within a selected time period, e.g., 24 hours.In certain embodiments, the PI3K gamma inhibitor or a compound asdescribed herein (e.g., a compound of any of Formulae (I″), (I′), (A′),(I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI), (XII),(XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound 4, as describedherein) is administered at a dose such that the compound's level in thesubject's blood does not rise higher than a predetermined level, e.g.,the IC20 of PI3K-delta, within a selected time period, e.g., 24 hours.

In some embodiments of the methods or uses disclosed herein, the subjectis refractory or has failed to respond to a PI3K-δ inhibitor. In someembodiments, the subject is naive to immunotherapy treatment. In someembodiments, the subject is or has been responsive to an immunotherapytreatment. In one embodiment, the immunotherapy treatment is a treatmentwith a PD-1 or PD-L1 inhibitor.

In other embodiments, a method of reducing CXCL12-induced CD3+ T cellmigration, or CXCL12-induced differentiated macrophage migration into atumor microenvironment, in a subject is provided. The method includesadministering to the subject a PI3K gamma inhibitor or a compound asdescribed herein (e.g., a compound of any of Formulae (I″), (I′), (A′),(I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI), (XII),(XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound 4, as describedherein), in an amount sufficient to reduce or inhibit the CXCL12-inducedCD3+ T cell migration, or CXCL12-induced differentiated macrophagemigration into a tumor microenvironment in the subject.

In some embodiments of the methods or uses disclosed herein, the subjecthas, or is identified as having, a reduction in p-AKT levels afteradministration of the PI3K gamma inhibitor or a compound as describedherein.

In some embodiments of the methods or uses disclosed herein, the PI3Kgamma inhibitor is administered at a dose such that it selectivelyinhibits PI3K-gamma but achieves less than 10% or 20% inhibition ofPI3K-delta.

In some embodiments of the methods or uses disclosed herein, the PI3Kgamma inhibitor or a compound as described herein is administered at adose such that the compound's level in the subject's blood does not risehigher than a predetermined level, e.g., the IC50 of PI3K-delta, withina selected time period, e.g., 24 hours. In some embodiments of themethods or uses disclosed herein, the PI3K gamma inhibitor or a compoundas described herein is administered at a dose such that the compound'slevel in the subject's blood does not rise higher than a predeterminedlevel, e.g., the IC20 of PI3K-delta, within a selected time period,e.g., 24 hours.

In some embodiments, a method of reducing one or more activities of apro-tumor immune cell in a subject having a cancer is provided. Themethod includes administering to the subject a PI3K gamma inhibitor or acompound as described herein (e.g., a compound of any of Formulae (I″),(I′), (A′), (I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X),(XI), (XII), (XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound 4,as described herein), in an amount sufficient to reduce or inhibit theone or more activities of the pro-tumor immune cell.

In some embodiments, the pro-tumor immune cell is a T-cell, an M2macrophage, a stromal cell, a dendritic cell, an endothelial cell, or amyeloid cell. In one embodiment, the myeloid cell is a tumor associatedsuppressive myeloid cell. In one embodiment, the tumor associatedsuppressive myeloid cell is a tumor associated macrophage (TAM), amyeloid derived suppressor cell (MDSC), a monocytic immature myeloidcell (iMc), or a granulocytic iMc/neutrophil.

In certain embodiments, the subject has, or is identified as having, adecrease in numbers of pro-tumor immune cells in a tumormicroenvironment, compared to a reference value, after administration ofthe PI3K gamma inhibitor or a compound as described herein.

In certain embodiments, the amount of the administered is sufficient toproduce a decrease in numbers of pro-tumor immune cells in a tumormicroenvironment, compared to a reference value, after administration ofthe PI3K gamma inhibitor or a compound as described herein.

In certain embodiments, the subject has, or is identified as having,increased activity of anti-tumor immune cells, compared to a referencevalue, after administration of the PI3K gamma inhibitor or a compound asdescribed herein.

In certain embodiments, the amount of the PI3K gamma inhibitor or acompound as described herein is sufficient to produce increased activityof anti-tumor immune cells, compared to a reference value, afteradministration of the PI3K gamma inhibitor or the compound as describedherein.

In certain embodiments, the subject has, or is identified as having,increased infiltration of anti-tumor immune cells into a tumormicroenvironment, compared to a reference value, after administration ofthe PI3K gamma inhibitor or a compound as described herein.

In certain embodiments, the amount of PI3K-gamma inhibitor is sufficientto produce increased infiltration of anti-tumor immune cells into atumor microenvironment, compared to a reference value, afteradministration of the PI3K gamma inhibitor or a compound as describedherein.

In certain embodiments, the subject has, or is identified as having, anincrease in number of anti-tumor immune cells in a tumormicroenvironment, compared to a reference value, after administration ofthe PI3K gamma inhibitor or a compound as described herein.

In certain embodiments, the amount of PI3K-gamma inhibitor is sufficientto produce an increase in number of anti-tumor immune cells in a tumormicroenvironment, compared to a reference value, after administration ofthe PI3K gamma inhibitor or a compound as described herein.

In certain embodiments, the cancer is a CLL. In some embodiments, thetumor microenvironment is a CLL proliferation center.

In certain embodiments, the subject has, or is identified as having,reduced tumor volume, compared to a reference value, afteradministration of the PI3K gamma inhibitor or a compound as describedherein.

In certain embodiments, the amount of the PI3K gamma inhibitor or acompound as described herein is sufficient to produce reduced tumorvolume, compared to a reference value, after administration of the PI3Kgamma inhibitor or the compound as described herein.

In certain embodiments, the amount of the PI3K gamma inhibitor or acompound as described herein is sufficient to produce a reduction of atleast 10%, 20%, 30%, 50%, 60%, or 60% in tumor volume, compared to areference value, after administration of the PI3K gamma inhibitor or thecompound as described herein.

In certain embodiments, the subject has, or is identified as having, anincreased level of apoptosis in the cancer cells, compared to areference value, after administration of the PI3K gamma inhibitor or acompound as described herein.

In certain embodiments, the amount of PI3K gamma inhibitor is sufficientto produce an increased level of apoptosis in the cancer cells, comparedto a reference value, after administration of the PI3K gamma inhibitoror a compound as described herein.

In certain embodiments, the subject has, or is identified as having, a10%, 20%, 30%, 40%, or 50% increase in apoptosis in the cancer cells,compared to a reference value, after administration of the PI3K gammainhibitor or a compound as described herein.

In certain embodiments, the amount of the PI3K gamma inhibitor or acompound as described herein is sufficient to produce a 10%, 20%, 30%,40%, or 50% increase in apoptosis in the cancer cells, compared to areference value, after administration of the PI3K gamma inhibitor or thecompound as described herein.

In certain embodiments, the anti-tumor immune cell is an M1 macrophage.

In certain embodiments, the one activity is choosen from one or more ofmigration of the cell, or signaling to an anti-tumor immune cell.

In certain embodiments, the subject has, or is determined to havereduced levels of p-AKT in the pro-tumor immune cell, compared to areference value, after administration of the PI3K gamma inhibitor or thecompound.

In certain embodiments, the amount is sufficient to reduce p-AKT in thepro-tumor immune cell, compared to a reference value, afteradministration of the PI3K gamma inhibitor or the compound.

In certain embodiments, the subject has, or is determined to have areduction of p-AKT levels by about 10%, 20%, 30%, 40%, 50%, or 60%,compared to a reference value, after administration of the PI3K gammainhibitor or the compound.

In certain embodiments, the subject has, or is determined to have areduction of p-AKT levels by about 10%, 20%, 30%, 40%, 50%, or 60%,compared to a reference value, after administration of the PI3K gammainhibitor or the compound.

In certain embodiments, the subject has, or is determined to have lungcancer, breast cancer, colon cancer, or glioblastoma. In certainembodiments, the breast cancer is triple negative breast cancer.

In some embodiments, a method of reducing the level of M2 macrophages ina tumor microenvironment in a subject having a cancer is provided. Themethod includes administering to the subject a PI3K gamma inhibitor or acompound as described herein (e.g., a compound of any of Formulae (I″),(I′), (A′), (I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X),(XI), (XII), (XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound 4,as described herein), in an amount sufficient to reduce the level of M2macrophages in a tumor microenvironment.

In certain embodiments, reducing the level of M2 macrophages comprisesreducing or inhibiting the differentiation of a tumor associated myeloidcell into an M2 macrophage. Differentiation into an M2 macrophage can bemeasured by decreased ARG1 levels compared to a reference value, afteradministration of the compound.

In certain embodiments, the ARG1 level is reduced by at least 10%, 20%,30%, 40%, 50%, 60%, 70%, 80%, or 90% compared to a reference value,after administration of the compound.

In certain embodiments, differentiation into an M2 macrophage ismeasured by decreased VEGF levels compared to a reference value, afteradministration of the compound.

In certain embodiments, the VEGF level is reduced by at least 10%, 20%,30%, 40%, 50%, 60%, 70%, 80%, or 90% compared to a reference value,after administration of the compound.

In certain embodiments, the subject has, or is determined to have, anormal level of differentiation of myeloid cells into M1 macrophages.

In certain embodiments, the amount is such that the compound does notreduce differentiation of myeloid cells into M1 macrophages.

In certain embodiments, the subject has, or is determined to have,increased anti-tumor immune attack by effector T cells, reducedvascularization of a tumor, reduced ECM breakdown, decreased tumorgrowth, or any combination thereof, compared to a reference value, afteradministration of the compound.

In certain embodiments, the cancer is, or is determined to be, a solidtumor (e.g., a cancer chosen from lung cancer, breast cancer, coloncancer, or glioblastoma).

In certain embodiments, the cancer is, or is determined to be, ahematological cancer.

Additional Combinations Therapies:

In other embodiments, provided herein is a method of treating aPI3K-mediated disorder in a subject, comprising administering to asubject a PI3K gamma inhibitor or a compound as described herein (e.g.,a compound of any of Formulae (I″), (I′), (A′), (I), (A), (II), (III),(IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII), (XIV), (XV),(XVI), and (XVII), e.g., Compound 4, as described herein) and animmunomodulator.

In certain embodiments, the PI3K-mediated disorder is a cancer,autoimmune disease, or inflammatory disease.

In one embodiment, the cancer is of hematopoietic origin, e.g., a cancerchosen from a leukemia or lymphoma, e.g., B-cell lymphoma, T-celllymphoma, non-Hodgkin's lymphoma, Hodgkin lymphoma, or anaplastic largecell lymphoma. In one embodiment, the lymphoma is follicular B celllymphoma.

In other embodiments, the cancer is a solid tumor, e.g., a cancer chosenfrom a breast cancer, a lung cancer, a colon cancer, melanoma, or aglioblastoma. In one embodiment, the cancer is melanoma, bladder cancer,head and neck cancer, lung cancer (e.g., non-small cell lung cancer),renal cell carcinoma, ovarian cancer, breast cancer (e.g.,triple-negative breast cancer), colon cancer, or glioblastoma.

In some embodiments, the subject is naive to immunotherapy treatment. Insome embodiments, the subject is or has been responsive to animmunotherapy treatment. In one embodiment, the immunotherapy treatmentis a treatment with a PD-1 or PD-L1 inhibitor.

In one embodiment, the immunomodulator is an immune checkpoint therapy,e.g., an immune checkpoint therapy chosen from is an inhibitor of PD-1,PD-L1, PD-L2, CTLA4, TIM3, LAG3, VISTA, BTLA, TIGIT, LAIR1, CD160, 2B4,TGFR-beta, or IDO/TDO, or any combination thereof. In one embodiment,the immune checkpoint therapy is an inhibitor of CTLA-4, PD-1, or PD-L1.The immune checkpoint therapy can be chosen from an antibody or fragmentthereof, an inhibitory nucleic acid, a soluble ligand, or a fusion of animmune checkpoint therapy (e.g., CTLA-4, PD-1, or PD-1 ligand) with a Fcregion of an immunoglobulin.

In certain embodiments, the immunomodulator is an activator of acostimulatory molecule. In one embodiment, the agonist of thecostimulatory molecule is chosen from an agonist (e.g., an agonisticantibody or antigen-binding fragment thereof, or a soluble fusion) ofOX40, CD2, CD27, CDS, ICAM-1, LFA-1 (CD11a/CD18), ICOS (CD278), 4-1BB(CD137), GITR, CD30, CD40, BAFFR, HVEM, CD7, LIGHT, NKG2C, SLAMF7,NKp80, CD160, B7-H3 or CD83 ligand.

The certain embodiments, the immunomodulator is chosen from acostimulatory ligand, a MCSF/CSF-1R inhibitor, an immunostimulant, aCXCR4/CXCL12 inhibitor, a CCL2 inhibitor, or a CCR2 inhibitor.

In certain embodiments, the PI3K gamma inhibitor or a compound asdescribed herein and the immunomodulator are in a single dosage form.

In certain embodiments, the PI3K gamma inhibitor or a compound asdescribed herein and the immunomodulator are in separate dosage forms.

In certain embodiments, the PI3K gamma inhibitor or a compound asdescribed herein and the immunomodulator are administered concurrently.

In certain embodiments, the PI3K gamma inhibitor or a compound asdescribed herein is administered subsequent to the immunomodulator.

In certain embodiments, the PI3K gamma inhibitor or a compound asdescribed herein is administered prior to the immunomodulator.

In certain embodiments, the effective amount of the PI3K gamma inhibitoror a compound as described herein, the immunomodulator, or both that isan amount sufficient to cause a decrease in tumor growth of at least10%, 20%, 30%, 40%, or 50% compared to a reference value, is reduced.

In certain embodiments, the subject has, or is determined to have, adecrease in tumor growth of at least 10%, 20%, 30%, 40%, or 50% comparedto a reference value, after administration of the PI3K gamma inhibitoror the compound.

In certain embodiments, the method comprises administration an immunecheckpoint therapy, e.g., a PD-1 or PD-L1 inhibitor (e.g., an anti-PD-1antibody or an anti-PD-L1 antibody).

In certain embodiments, the method further comprises administering aneffective amount of a PI3K-δ inhibitor or the compound to the subject.

In certain embodiments, the level of the PI3K gamma inhibitor in thesubject's blood is above a predetermined value, e.g., the IC50 ofPI3K-gamma, during at least 70%, 80%, 90%, 95%, 97%, 98%, 99%, or 99% ofa selected time period, e.g., 24 hours, immediately following thetreatment.

In certain embodiments, the level of the PI3K gamma inhibitor in thesubject's blood is below a predetermined value, e.g., the IC50 ofPI3K-delta, during at least 70%, 80%, 90%, 95%, 97%, 98%, 99%, or 99% ofa selected time period, e.g., 24 hours, immediately following thetreatment. In certain embodiments, the level of the PI3K gamma inhibitoror the compound in the subject's blood is below a predetermined value,e.g., the IC20 of PI3K-delta, during at least 70%, 80%, 90%, 95%, 97%,98%, 99%, or 99% of a selected time period, e.g., 24 hours, immediatelyfollowing the treatment.

Compositions comprising the aforesaid combinations of PI3K gammainhibitor and the immunomodulator are also provided. The compositionscan be provided in the same or in separate dosage forms. Thecompositions can further include a pharmaceutically acceptableexcipient.

In other embodiments, the invention features a method of treating aPI3K-mediated disorder in a subject, comprising administering to thesubject a synergistic combination of a PI3K gamma inhibitor (e.g., acompound of any of Formulae (I″), (I′), (A′), (I), (A), (II), (III),(IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII), (XIV), (XV),(XVI), and (XVII), e.g., Compound 4, as described herein) and a PI3K-δinhibitor.

In certain embodiments, the PI3K-mediated disorder is a cancer,autoimmune disease, or inflammatory disease.

In one embodiment, the cancer is of hematopoietic origin, e.g., a cancerchosen from a leukemia or lymphoma, e.g., B-cell lymphoma, T-celllymphoma, non-Hodgkin's lymphoma, Hodgkin lymphoma, or anaplastic largecell lymphoma.

In other embodiments, the cancer is a solid tumor, e.g., a cancer chosenfrom a breast cancer, a lung cancer, a colon cancer, or a glioblastoma.

In certain embodiments, the PI3K gamma inhibitor and the PI3K-δinhibitor are in a single dosage form.

In certain embodiments, the PI3K gamma inhibitor and the PI3K-δinhibitor are in separate dosage forms.

In certain embodiments, the PI3K gamma inhibitor and the PI3K-δinhibitor are administered concurrently.

In certain embodiments, the PI3K gamma inhibitor is administeredsubsequent to the PI3K-δ inhibitor.

In certain embodiments, the PI3K gamma inhibitor is administered priorto the PI3K-δ inhibitor.

In certain embodiments, the PI3K gamma inhibitor is a selective PI3K-δinhibitor, e.g., has a delta/gamma selectivity ratio of greater thanabout 1 to <10, greater than about 10 to <50, or greater than about 50to <350.

In certain embodiments, the concentration of the PI3K gamma inhibitor,e.g., Compound 4, that is required to achieve inhibition, e.g., 50%inhibition, is lower (e.g., at least 20%, 30%, 40%, or 50% lower) whenthe PI3K gamma inhibitor is administered in combination with the PI3K-δinhibitor than when the PI3K gamma inhibitor is administeredindividually.

In certain embodiments, the concentration of the PI3K-δ inhibitor thatis required to achieve inhibition, e.g., 50% inhibition, is lower (e.g.,at least 20%, 30%, 40%, or 50% lower) when the second therapeutic agentis administered in combination with the PI3K gamma inhibitor than whenthe PI3K-δ inhibitor is administered individually.

In certain embodiments, the dose of the PI3K gamma inhibitor that isrequired to achieve inhibition, e.g., 50% inhibition, is lower (e.g., atleast 20%, 30%, 40%, or 50% lower) when the PI3K gamma inhibitor isadministered in combination with the PI3K-δ inhibitor than when the PI3Kgamma inhibitor is administered individually.

In certain embodiments, the dose of the second therapeutic agent that isrequired to achieve inhibition, e.g., 50% inhibition, is lower (e.g., atleast 20%, 30%, 40%, or 50% lower) when the second therapeutic agent isadministered in combination with the PI3K gamma inhibitor than when thePI3K-δ inhibitor is administered individually.

In certain embodiments, the combination is synergistic as indicated by acombination index value that is less than 0.7, 0.5, or 0.1 for thecombination of the PI3K gamma inhibitor and the PI3K-δ inhibitor.

In certain embodiments, the combination index value is assessed at 50%inhibition.

In certain embodiments, the combination index value is assessed at 50%growth inhibition.

In certain embodiments, the combination is synergistic as indicated by aSynergy Score that is greater than 1, 2, or 3, for the combination ofthe PI3K gamma inhibitor and the second therapeutic agent.

In certain embodiments, the combination is synergistic as indicated by aSynergy Score that is greater than 1, 2, or 3, for the combination ofthe PI3K gamma inhibitor and the PI3K-δ inhibitor for inhibition orgrowth inhibition.

In certain embodiments, the anti-cancer effect provided by thecomposition greater than the anti-cancer effect provided by themonotherapy with the PI3K gamma inhibitor or pharmaceutically acceptableform thereof, e.g., is at least 2 fold greater, at least 3 fold greater,at least 5 fold greater, or at least 10 fold greater.

In certain embodiments, the anti-cancer effect provided by thecomposition is greater than the anti-cancer effect provided by themonotherapy with the PI3K-δ inhibitor or pharmaceutically acceptableform thereof, e.g., is at least 2 fold greater, at least 3 fold greater,at least 5 fold greater, or at least 10 fold greater.

In certain embodiments, one or more side effects of the composition isreduced compared with the side effects of administering a monotherapycomprising the PI3K gamma inhibitor or pharmaceutically acceptable formthereof, without PI3K-δ inhibitor or pharmaceutically acceptable formthereof, at a dose that achieves the same therapeutic effect.

In certain embodiments, one or more side effects of the composition isreduced compared with the side effects of administering a monotherapycomprising the PI3K-δ inhibitor or pharmaceutically acceptable formthereof, without the PI3K gamma inhibitor or pharmaceutically acceptableform thereof, at a dose that achieves the same therapeutic effect.

In another aspect, the invention features a composition comprising asynergistic combination of a a PI3K gamma inhibitor (e.g., a compound ofany of Formulae (I″), (I′), (A′), (I), (A), (II), (III), (IV), (V),(VI), (VII), (IX), (X), (XI), (XII), (XIII), (XIV), (XV), (XVI), and(XVII), e.g., Compound 4, as described herein) and a PI3K-δ inhibitor.The compositions can be provided in the same or in separate dosageforms.

In certain embodiments, the PI3K-δ inhibitor is a selective PI3K-δinhibitor, e.g., has a delta/gamma selectivity ratio of greater thanabout 1 to <10, greater than about 10 to <50, or greater than about 50to <350.

In certain embodiments, the molar ratio of the PI3K gamma inhibitor tothe PI3K-δ inhibitor is in the range of from about 10000:1 to about1:10000.

In certain embodiments, the molar ratio of the PI3K gamma inhibitor tothe PI3K-δ inhibitor is in the range of from about 10:1 to about 1:10.

In certain embodiments, the composition comprises the PI3K gammainhibitor, or pharmaceutically acceptable form thereof, at an amount ofin the range of from about 0.01 mg to about 75 mg and the PI3K-δinhibitor, or pharmaceutically acceptable form thereof, at an amount ofin the range of from about 0.01 mg to about 1100 mg.

In certain embodiments, the PI3K gamma inhibitor or pharmaceuticallyacceptable form thereof, and the PI3K-δ inhibitor or pharmaceuticallyacceptable form thereof, are the only therapeutically activeingredients.

In certain embodiments, the PI3K gamma inhibitor or pharmaceuticallyacceptable form thereof, and the PI3K-δ inhibitor or pharmaceuticallyacceptable form thereof, are in a single dosage form.

In certain embodiments, the PI3K gamma inhibitor or pharmaceuticallyacceptable form thereof, and the PI3K-δ inhibitor or pharmaceuticallyacceptable form thereof, are in separate dosage forms.

In certain embodiments, the combination of the PI3K gamma inhibitor andthe PI3K-δ inhibitor are synergistic in treating a cancer, inflammatorydisease, or autoimmune disease.

In certain embodiments, the concentration of the PI3K gamma inhibitorthat is required to achieve inhibition, e.g., 50% inhibition, is lower(e.g., at least 20%, 30%, 40%, or 50% lower) when the Compound 4 isadministered in combination with the PI3K-δ inhibitor than when theCompound 4 is administered individually.

In certain embodiments, the concentration of the second therapeuticagent that is required to achieve inhibition, e.g., 50% inhibition, islower (e.g., at least 20%, 30%, 40%, or 50% lower) when the secondtherapeutic agent is administered in combination with the PI3K gammainhibitor than when the second therapeutic agent is administeredindividually.

In certain embodiments, the dose of the PI3K gamma inhibitor that isrequired to achieve inhibition, e.g., 50% inhibition, is lower (e.g., atleast 20%, 30%, 40%, or 50% lower) when the PI3K gamma inhibitor isadministered in combination with the PI3K-δ inhibitor than when the PI3Kgamma inhibitor is administered individually.

In certain embodiments, the dose of the second therapeutic agent that isrequired to achieve inhibition, e.g., 50% inhibition, is lower (e.g., atleast 20%, 30%, 40%, or 50% lower) when the second PI3K-δ inhibitor isadministered in combination with the PI3K gamma inhibitor than when thePI3K-δ inhibitor is administered individually.

In certain embodiments, wherein the combination is synergistic asindicated by a combination index value that is less than 0.7, 0.5, or0.1 for the combination of the PI3K gamma inhibitor and the PI3K-δinhibitor.

In certain embodiments, the combination index value is assessed at 50%inhibition.

In certain embodiments, the combination index value is assessed at 50%growth inhibition.

In certain embodiments, the combination is synergistic as indicated by aSynergy Score that is greater than 1, 2, or 3, for the combination ofthe PI3K gamma inhibitor and the PI3K-δ inhibitor

In certain embodiments, the combination is synergistic as indicated by aSynergy Score that is greater than 1, 2, or 3, for the combination ofthe PI3K gamma inhibitor and the PI3K-δ inhibitor for inhibition orgrowth inhibition.

In certain embodiments, the anti-cancer effect provided by thecomposition greater than the anti-cancer effect provided by themonotherapy with the PI3K gamma inhibitor or pharmaceutically acceptableform thereof, e.g., is at least 2 fold greater, at least 3 fold greater,at least 5 fold greater, or at least 10 fold greater.

In certain embodiments, the anti-cancer effect provided by thecomposition is greater than the anti-cancer effect provided by themonotherapy with the PI3K-δ inhibitor or pharmaceutically acceptableform thereof, e.g., is at least 2 fold greater, at least 3 fold greater,at least 5 fold greater, or at least 10 fold greater.

In certain embodiments, one or more side effects of the composition isreduced compared with the side effects of administering a monotherapycomprising the PI3K gamma inhibitor or the compound, or pharmaceuticallyacceptable form thereof, without the PI3K-δ inhibitor or the compound,pharmaceutically acceptable form thereof, at a dose that achieves thesame therapeutic effect.

In certain embodiments, one or more side effects of the composition isreduced compared with the side effects of administering a monotherapycomprising the PI3K-δ inhibitor or pharmaceutically acceptable formthereof, without the PI3K gamma inhibitor or pharmaceutically acceptableform thereof, at a dose that achieves the same therapeutic effect.

In certain embodiments, the compositions further comprise apharmaceutically acceptable excipient.

Bone Disorders

In another aspect, the invention features a method of treating orpreventing a bone disorder, e.g., a method of reducing osteoclastactivity in a subject having a bone formation disorder. The methodincludes administering a PI3K gamma inhibitor or a compound as describedherein (e.g., a compound of any of Formulae (I″), (I′), (A′), (I), (A),(II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII),(XIV), (XV), (XVI), and (XVII), e.g., Compound 4, as described herein)in an amount sufficient to treat or prevent the bone disorder.

In certain embodiments, the effective amount of the PI3K gamma inhibitoris an amount sufficient to reduce the number of osteoclasts in thesubject compared to a reference value, after administration of thecompound.

In certain embodiments, the effective amount of the PI3K gamma inhibitoris an amount sufficient to reduce the number of osteoclasts in thesubject by about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% comparedto a reference value, after administration of the compound.

In certain embodiments, the subject has, or is determined to have, areduction in the number of osteoclasts compared to a reference value,after administration of the compound.

In certain embodiments, the subject has, or is determined to have, areduction in the number of osteoclasts by about 10%, 20%, 30%, 40%, 50%,60%, 70%, 80%, or 90% compared to a reference value, afteradministration of the compound.

In certain embodiments, the effective amount is an amount sufficient toresult in decreased differentiation of a macrophage into an osteoclast.

In certain embodiments, the effective amount is an amount sufficient toresult in differentiation of macrophages into osteoclasts being reducedby about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90%.

In certain embodiments, the subject has, or is determined to have,decreased differentiation of a macrophage into an osteoclast.

In certain embodiments, the subject has, or is determined to have,differentiation of macrophages into osteoclasts being reduced by about10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90%.

In certain embodiments, differentiation is measured by identifying TRAPpositive cells.

In certain embodiments, which does not result in disruption of thefunction of a differentiated osteoclast.

In certain embodiments, the bone formation disorder is osteoporosis,osteoporosis pseudoglioma, hyperparathyroidism, hyperthyroidism, Paget'sdisease, hyperphosphatasemia, fibrous dysplasia, osteogenesisimperfecta, primary and secondary hyperparathyroidism and associatedsyndromes, hypercalciuria, metastatic osteolytic carcinomas (includingmedullary carcinoma of the thyroid gland, prostate, renal, breast, andlung cancer), or osteomalacia. In certain embodiments, the compoundsprovide herein is used in the treatment of osteolytic cancer metastases,fracture repair, rheumatoid arthritis, or other autoimmune diseases todamage of bone and joints.

In certain embodiments, provided herein are methods of treating orpreventing a bone formation disorder in a subject, comprisingadministering to the subject a therapeutically effective amount of acompound provided herein (e.g., a compound of any of Formulae (I″),(I′), (A′), (I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X),(XI), (XII), (XIII), (XIV), (XV), (XVI), and (XVII), e.g., a selectivePI3K-γ inhibitor, e.g., Compound 4), or a pharmaceutically acceptableform thereof.

In one embodiment, the number of osteoclasts in the subject is reducedby about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% compared to areference value, after administration of the compound. In oneembodiment, the differentiation of macrophage cells into osteoclasts isreduced by about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% comparedto a reference value, after administration of the compound. In oneembodiment, the differentiation of macrophage cells into osteoclasts ismeasured by identifying TRAP positive cells.

In one embodiment, the method does not result in disruption of thefunction of differentiated osteoclasts.

In one embodiment, the bone formation disorder is osteoporosis,osteoporosis pseudoglioma, hyperparathyroidism, hyperthyroidism, Paget'sdisease, hyperphosphatasemia, fibrous dysplasia, osteogenesisimperfecta, primary and secondary hyperparathyroidism and associatedsyndromes, hypercalciuria, metastatic osteolytic carcinomas (includingmedullary carcinoma of the thyroid gland, prostate, renal, breast, andlung cancer), or osteomalacia.

Combination with Radiation Therapy:

In another aspect, the invention features a method of treating a solidtumor, comprising administering to a subject in need thereof aneffective amount of a PI3K gamma inhibitor or a compound as describedherein (e.g., a compound of any of Formulae (I″), (I′), (A′), (I), (A),(II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII),(XIV), (XV), (XVI), and (XVII), e.g., Compound 4, as described herein):

or a pharmaceutically acceptable form thereof.

In embodiment, the treatment can be, e.g., either prior to administeringthe radiation therapy, after administering radiation therapy, or at thesame time as administering radiation therapy. In another embodiment, thetreatment can be, e.g., either after administering radiation therapy, orat the same time as administering radiation therapy. In anotherembodiment, the treatment can be after administering radiation therapy.

In certain embodiments, the cancer is selected from one or more of: acancer of the pulmonary system, a brain cancer, a cancer of thegastrointestinal tract, a skin cancer, a genitourinary cancer, apancreatic cancer, a lung cancer, a medullobastoma, a basal cellcarcinoma, a glioma, a breast cancer, a prostate cancer, a testicularcancer, an esophageal cancer, a hepatocellular cancer, a gastric cancer,a gastrointestinal stromal tumor (GIST), a colon cancer, a colorectalcancer, an ovarian cancer, a melanoma, a neuroectodermal tumor, head andneck cancer, a sarcoma, a soft-tissue sarcoma, fibrosarcoma,myxosarcoma, liposarcoma, a chondrosarcoma, an osteogenic sarcoma, achordoma, an angiosarcoma, an endotheliosarcoma, a lymphangiosarcoma, alymphangioendotheliosarcoma, a synovioma, a mesothelioma, aleiomyosarcoma, a cervical cancer, a uterine cancer, an endometrialcancer, a carcinoma, a bladder carcinoma, an epithelial carcinoma, asquamous cell carcinoma, an adenocarcinoma, a bronchogenic carcinoma, arenal cell carcinoma, a hepatoma, a bile duct carcinoma, aneuroendocrine cancer, a carcinoid tumor, diffuse type giant cell tumor,and glioblastoma. In certain embodiments, the solid tumor is a lungtumor, breast tumor, colon tumor, brain tumor, bone tumor, glioblastoma,or a metastatic lesion thereof. In one embodiment, the combination ofradiation and/or PI3K-gamma inhibition is such that accumulation oftumor supporting-myeloid cells into the radiated tumor is reduced orprevented, thus impairing tumor regrowth following radiation therapy.

Intratracheal Administration

In some aspects, the present disclosure provides a method of treating(e.g., ameliorating, preventing, and/or managing) a pulmonary orrespiratory disease in a subject, comprising administering to a subjectin need thereof by inhalation a therapeutically or prophylacticallyeffective amount of a PI3K gamma inhibitor or a compound as describedherein (e.g., a compound of any of Formulae (I″), (I′), (A′), (I), (A),(II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII),(XIV), (XV), (XVI), and (XVII), e.g., Compound 4, as described herein):

or an enantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof.

In some aspects, this disclosure provides a method of eliciting aprolonged anti-inflammatory effect in a lung in a subject suffering froma pulmonary or respiratory disease, comprising administering to thesubject by inhalation a therapeutically or prophylactically effectiveamount of the PI3K gamma inhibitor or a compound as described herein(e.g., Compound 4) or an enantiomer, a mixture of enantiomers, or amixture of two or more diastereomers thereof, or a pharmaceuticallyacceptable form thereof, wherein the compound is retained in lung for aperiod longer than what is provided by oral administration.

In some embodiments, the compound is retained in the lung for a periodthat is sufficient to administer the compound once a day, twice a day,three times a day, four times a day, five times a day, or once every twoday.

In some embodiments, the compound is retained in lung for about 1 hour,about 3 hours, about 6 hours, about 12 hours, about 24 hours, about 48hours, or about 72 hours longer than what is provided by oraladministration.

In some embodiments, the compound is administered at a dose of less than0.01 μg/kg/day, less than 0.02 μg/kg/day, less than 0.05 μg/kg/day, lessthan 0.1 μg/kg/day, less than 0.2 μg/kg/day, less than 0.5 μg/kg/day,less than 1 μg/kg/day, less than 2 μg/kg/day, less than 5 μg/kg/day,less than 10 μg/kg/day, less than 20 μg/kg/day, less than 50 μg/kg/day,or less than 100 μg/kg/day. In some embodiments, the compound isadministered at a dose of about 0.01 μg/kg/day, about 0.02 μg/kg/day,about 0.05 μg/kg/day, about 0.1 μg/kg/day, about 0.2 μg/kg/day, about0.5 μg/kg/day, about 1 μg/kg/day, about 2 μg/kg/day, about 5 μg/kg/day,about 10 μg/kg/day, about 20 μg/kg/day, about 50 μg/kg/day, or about 100μg/kg/day. In some embodiments, the compound is administered at a doseof from about 0.01 μg/kg/day to about 100 μg/kg/day, from about 0.01μg/kg/day to about 50 μg/kg/day, from about 0.01 μg/kg/day to about 20μg/kg/day, from about 0.01 μg/kg/day to about 10 μg/kg/day, from about0.01 μg/kg/day to about 5 μg/kg/day, from about 0.01 μg/kg/day to about1 μg/kg/day, from about 0.05 μg/kg/day to about 1 μg/kg/day, or fromabout 0.1 μg/kg/day to about 1 μg/kg/day.

In some embodiments, the compound is administered once daily (QD), twicedaily (BID), three times daily (TID), or four times daily (QID).

In some embodiments, administering an effective amount of the compounddoes not result in, or results in reduced, one or more common sideeffects associated with treatment of pulmonary or respiratory diseases.In some embodiments, the common side effect associated with treatment ofpulmonary or respiratory diseases is oral candidiasis, thrush,dysphonia, reflex cough, bronchospasm, poor growth, decreased bonedensity, disseminated varicella infection, easy bruising, cataracts,glaucoma, adrenal gland suppression, stomach upset, headache, liver testabnormalities, skin rashes, Churg Strauss syndrome, bad taste in month,cough, itching, sore throat, sneezing, stuffy nose, shortness of breath,wheezing, viral illness, upper respiratory tract infections, sinusitis,feeling dizzy or faint, hives, changes in voice, swelling of the tongue,or difficulty in swallowing.

In some embodiments, administering an effective amount of the compoundreduces one of more of symptoms associated with pulmonary or respiratorydiseases. In some embodiments, the symptom associated with pulmonary orrespiratory diseases is wheezing, coughing, chest tightness, shortnessof breath, difficulty in breathing, or use of accessory muscle.

In some embodiments, administering an effective amount of the compoundby inhalation results in higher than about 20%, higher than about 30%,higher than about 40%, or higher than about 50% of the administered doseof the compound remaining in lung of the subject at about 24 hours afterthe administration.

In some embodiments, administering an effective amount of the compoundby inhalation results in that the lung concentration of the compound isabout 100, about 200, about 500, about 1000, about 2000, about 3000,about 4000, about 5000, about 6000, about 7000, about 8000, about 9000,or about 10000 times higher than the plasma concentration of thecompound at about 5 hours after the administration. In some embodiments,administering an effective amount of the compound by inhalation resultsin that the lung concentration of the compound is about 100, about 200,about 500, about 1000, about 2000, about 3000, about 4000, about 5000,about 6000, about 7000, about 8000, about 9000, or about 10000 timeshigher than the plasma concentration of the compound at about 12 hoursafter the administration. In some embodiments, administering aneffective amount of the compound by inhalation results in that the lungconcentration of the compound is about 100, about 200, about 500, about1000, about 2000, about 3000, about 4000, about 5000, about 6000, about7000, about 8000, about 9000, or about 10000 times higher than theplasma concentration of the compound at about 24 hours after theadministration.

In some embodiments, the pulmonary or respiratory disease is selectedfrom the group consisting of pulmonary inflammation, asthma, cysticfibrosis, emphysema, chronic obstructive pulmonary disorder (COPD),chronic bronchitis, bronchiectasis, acute respiratory distress syndrome,restrictive lung diseases, respiratory tract infections, pleural cavitydiseases, pulmonary vascular disease, pulmonary embolism, pulmonaryarterial hypertension, pulmonary edema, pulmonary hemorrhage, andpulmonary hyperplasia.

In some embodiments, the pulmonary or respiratory disease is chronicobstructive pulmonary disorder. In some embodiments, the pulmonary orrespiratory disease is asthma. In some embodiments, the asthma isselected from the group consisting of severe or refractory asthma,atopic asthma, non-atopic asthma, type 1 brittle asthma, type 2 brittleasthma, asthma attack, status asthmaticus, exercise-induced asthma, andoccupational asthma.

In some embodiments, the subject is a mammal. In some embodiments, thesubject is a human.

In some embodiments, the method further comprises administration of anadditional therapeutic agent, e.g., an agent selected from one or moreof Arcapta (indacaterol maleate inhalation powder), Daliresp(roflumilast), Dulera (mometasone furoate+formoterol fumaratedihydrate), Alvesco (ciclesonide), Brovana (arformoterol tartrate),Spiriva HandiHaler (tiotropium bromide), Xolair (omalizumab), Qvar(beclomethasone dipropionate), Xopenex (levalbuterol), DuoNeb (albuterolsulfate and ipratropium bromide), Foradil Aerolizer (formoterol fumarateinhalation powder), Accolate (zafirlukast), Singulair (montelukastsodium), Flovent Rotadisk (Rotadisk (fluticasone propionate inhalationpowder), Tilade (nedocromil sodium), Vanceril (beclomethasonedipropionate, 84 mcg), Zyflo (Zileuton), and Azmacort (triamcinoloneacetonide) Inhalation Aerosol.

Abbreviations for PI3K isoforms are provided interchangeably throughoutas either the Greek letter or corresponding name. For example, the termsPI3K-γ and PI3K-gamma, or PI3K-δ and PI3K-delta are used interchangeablythroughout.

INCORPORATION BY REFERENCE

All publications, patents, and patent applications mentioned in thisspecification are herein incorporated by reference to the same extent asif each individual publication, patent, or patent application wasspecifically and individually indicated to be incorporated by reference.In case of conflict, the present application, including any definitionsherein, will control.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows the effect of Compound BB in the collagen induced arthritisrat model as measured by the mean ankle diameter over time.

FIG. 2 shows the effect of Compound BB in the collagen induced arthritisrat model as measured by the individual histopathology scores forinflammation, pannus, cartilage damage, bone resorption, and periostealbone formation.

FIG. 3 shows the effect of Compound BB in the collagen induced arthritisrat model as measured by periosteal bone measure.

FIG. 4 shows the effect of Compound BB on IL-8 induced neutrophilmigration in a mouse air pouch model.

FIG. 5 shows the effect of Compound AA on IL-8 induced neutrophilmigration in a mouse air pouch model.

FIG. 6 shows an isobologram depicting the synergistic effect of thecombination of a PI3K delta-selective compound, Compound AA, and a PI3Kgamma selective compound, Compound BB, in TMD8 cell line.

FIG. 7 shows an isobologram depicting the synergistic effect of thecombination of a PI3K delta-selective compound, Compound AA and a PI3Kgamma selective compound, Compound BB, in Farage cell line.

FIG. 8 shows the effects of a PI3K delta selective compound, CompoundAA, and a PI3K selective compound, Compound BB, on migration of chroniclymphocytic leukemia (CLL) associated peripheral T-cells.

FIG. 9 shows that CXCL12 induced pAKT in T-cells is PI3K-γ dependent.

FIG. 10 is a graph showing the total cell counts per milliliter in abronchoalveolar lavage (BAL) sample when the lipopolysaccharide (LPS)induced inflammatory rat model is exposed to Compound AA or Compound BB.

FIG. 11 is a graph showing the neutrophil counts per milliliter in abronchoalveolar lavage (BAL) sample when the lipopolysaccharide (LPS)induced inflammatory rat model is exposed to Compound AA or Compound BB.

FIG. 12 is a graph showing TNFα pictogram per milliliter in abronchoalveolar lavage (BAL) sample when the lipopolysaccharide (LPS)induced inflammatory rat model is exposed to Compound AA or Compound BB.

FIG. 13 shows the effects of Compound AA and Compound BB on CXCL12(SDF-1a) induced p-AKT in M2 phenotype macrophage.

FIG. 14 shows that co-cultures of M2 macrophages with CLL cells led toextended CLL cell survival.

FIG. 15 is a schematic of the differentiation of myeloid progenitorcells and interactions between certain T cells.

FIG. 16 illustrates differentiation of a myeloid cell into an M1macrophage or M2 macrophage.

FIG. 17 shows that Compound BB prevents RANKL driven differentiation ofosteoclasts from bone marrow macrophages.

FIG. 18 shows the mean unbound concentration of Compound BB in plasma, 1or 5 hours after dosing.

FIG. 19 shows a 24-hour time course of unbound plasma concentration ofCompound BB. The five horizontal lines represent the IC₅₀s (and oneIC₉₀) of four PI3K isoforms. From top to bottom, the horizontal linescorrespond to the IC₅₀ for PI3K-α, IC₅₀ for PI3K-β, IC₅₀ for PI3K-δ,IC₉₀ for PI3K-γ, and IC₅₀ for PI3K-γ.

FIG. 20 shows the mean plasma unbound Compound 88 concentration-timeprofiles for female C57 mice administered 2, 7.5 or 15 mg/kg PO.

FIG. 21 shows that Compound AA is a highly selective inhibitor of IFN-γin a Concanavalin A-induced model for activation of human T cells.Compound BB has little to no effect in this assay at isoform selectiveconcentrations.

FIGS. 22A and 22B show the level of ARG1 (Arginase-1) in murinemacrophages polarized to M2 phenotype in response to different doses ofCompound BB and Compound AA, respectively.

FIG. 23 shows that CLL cell survival is enhanced upon M2 macrophageco-culture. The x-axis corresponds to time, and the y-axis correspondsto % CLL viability.

FIGS. 24A and 24B show that Compound BB suppresses VEGF and ARG1 in anMyeloid Derived Suppressor Cell differentiation assay, respectively.

FIGS. 25A and 25B show that Compound BB demonstrates single agentactivity in the Lewis Lung Carcinoma model.

FIG. 26. shows the mean unbound Compound BB concentrations in tumor andplasma following 12 days of daily oral administration in NMP solution.

FIGS. 27A and 27B show the effect of Compound BB administered togetherwith an anti-PD-L1 checkpoint inhibitor in the CT26 colon cancer model.FIG. 27A shows the effect on tumor volume over time. FIG. 27B shows thepercent survival over time.

FIG. 28 shows that MDSC expansion after cyclophosphamide treatment isdecreased by Compound BB in the CT26 colon cancer model.

FIGS. 29A and 29B show the efficacy of Compound BB alone and incombination with 5-FU, respectively, in the MC38 colon cancer model. Thex-axis represents time and the y-axis represents tumor size in mm³.

FIG. 30 shows CD3 IHC analysis after treatment with Compound BB in MC38tumors.

FIG. 31A shows the CD3 and tumor volume correlation for vehicle. FIG.31B shows the CD3 and tumor volume correlation for Compound BB.

FIGS. 32A and 32B show the effects of Compound BB and/or Compound AA inthe DoHH2 human follicular B cell lymphoma model. FIG. 32B is a repeatstudy.

FIG. 33A shows reduced tumor growth with Compound BB and/or PDL-1inhibition in orthotopic 4T1 breast cancer model.

FIG. 33B shows reduced total luciferase flux with Compound BB and/orPDL-1 inhibition in orthotopic 4T1 breast cancer model.

FIG. 34 shows that Compound BB demonstrates single agent activity insubcutaneous 4T1 breast cancer model.

FIG. 35 shows that Compound BB alone achieves a reduction in tumorgrowth, while co-administration of Compound BB and 5-FU achieves afurther reduction.

FIG. 36 shows that Compound BB demonstrated single agent activity insubcutaneous 4T1 breast cancer model, while co-administration withdocetaxel, paclitaxel, or 5-FU achieved a further reduction in tumorgrowth.

FIGS. 37A, 37B, 37C, 37D, 37E, 37F, 37G, and 37H show CD11b+ or CD68+myeloid cell quantities in cancer tissue from Compound BB-treated orcontrol mice.

FIGS. 38A and 38B show the levels of different immune cells in treatedand untreated tumors. Specifically, FIG. 38A shows that TAMs aredecreased in Compound BB-treated LLC tumor brei model. FIG. 38B showsthat Monocytic iMCs are decreased in Compound BB-treated CT26subcutaneous tumors.

FIG. 39 shows that Compound BB reduces CD206+M2 polarized suppressivemacrophages in LLC tumor brei model.

FIG. 40 shows normalization to β-actin gives stable expression of GAPDHand 18S genes.

FIG. 41 shows the effect of Compound BB on markers Cd8b1, Pdcd1/PD1,Cd4, Cd3e, Foxp3, and C11a4 with or without PDL-1 antibody.

FIG. 42A shows M2 macrophage markers VEGF and ARG-1 are reduced byCompound BB with or without PDL-1 antibody. FIG. 42B shows the effect ofCompound BB with or without PDL-1 antibody on M2 macrophage markersARG1, VEGFa and ADM.

FIG. 43 shows that Compound BB oral exposure is high in the rat, dog andmonkey.

FIG. 44 shows that Compound BB exhibits dose-exposure proportionality.

FIG. 45 shows the concentrations of unbound Compound BB in a toxicitystudy.

FIG. 46 shows the survival of primary human CLL cells with and withoutco-cultured stromal cells.

FIG. 47A and FIG. 47B show the effects of Compound BB QD on tumor volumefrom day 9 to day 20 after implant and total luciferase flux in theGL-261 syngeneic glioma model. FIG. 47C is a repeat study that shows theeffects of Compound BB QD on tumor volume from day 9 to day 25 afterimplant.

FIG. 48 shows tumor growth after treatment with CompoundBB+/−cyclophosphamide in the LLC tumor brei model.

FIGS. 49A, 49B, 49C, 49D, and 49E show CD11b/Gr-1 plots after treatmentwith Compound BB+/−cyclophosphamide in the LLC tumor brei model. FIG.49A shows the percentage of cells that are CD3 relative to CD45+. FIG.49B shows the percentage of cells that are CD11b+Gr-1− relative toCD45+. FIG. 49C shows the percentage of cells that are CD11b+Gr-1+relative to CD45+.

FIG. 49D shows the percentage of cells that are CD11b-Gr-1− relative toCD45+. FIG. 49E shows the percentage of cells that are CD11b-Gr-1+relative to CD45+.

FIGS. 50A, 50B, 50C, and 50D show MDSC panel CD11b/Ly6C/Ly6G aftertreatment with Compound BB+/−cyclophosphamide in the LLC tumor breimodel. FIG. 50A shows the percentage of cells that are CD11b+ relativeto CD45. FIG. 50B shows the percentage of cells that are CD11b+Ly6C+relative to CD45+. FIG. 50C shows the percentage of cells that areCD11b+Ly6C+Ly6G+ relative to CD45+. FIG. 50D shows the percentage ofcells that are CD11b+Ly6C-Ly6G− relative to CD45+.

FIGS. 51A, 51B, 51C, 51D, and 51E show CD3/CD4/CD8 plots after treatmentwith Compound BB+/−cyclophosphamide in the LLC tumor brei model. FIG.51A shows the percentage of cells that are CD3+ relative to CD45+. FIG.51B shows the percentage of cells that are CD4+CD8− relative to CD45+.FIG. 51C shows the percentage of cells that are CD4+CD8+ relative toCD45+. FIG. 51D shows the percentage of cells that are CD4-CD8+ relativeto CD45+. FIG. 51E shows the percentage of cells that are CD4-CD8−relative to CD45+.

FIGS. 52A, 52B, 52C, 52D, and 52E show the percentage of cells that areCD45+, CD11b+, or CD206+, expressed as a percent of total cells, in LLCtumor brei model treated with Compound BB+/−cyclophosphamide. FIG. 52Ashows the percentage of cells that are CD45+ expressed as a percent oftotal cells. FIG. 52B shows the percentage of cells that are CD11bexpressed as a percent of total cells. FIG. 52C shows the percentage ofcells that are CD206 expressed as a percent of total cells. FIG. 52Dshows the percentage of cells that are CD11b expressed as a percent oftotal cells. FIG. 52E shows the percentage of cells that are CD206expressed as a percent of total cells.

FIGS. 53A, 53B, 53C, 53D, and 53E show the effect of Compound BBfollowing temozolomide treatment on GL-261 tumors.

FIG. 54A shows the effect of TMZ treatment versus vehicle. FIG. 54B showthe effects of Compound BB on tumor volume post TMZ treatment.

FIGS. 55A, 55B, and 55C show the effect of Compound BB and anti PDL-1 ontumor-derived cell mediated inhibition of T cell proliferation versusvehicle. FIG. 55A shows the effect of vehicle treated tumor-derived cellmediated T cell proliferation. FIG. 55B shows the effect of Compound BBtreated tumor-derived cell mediated T cell proliferation. FIG. 55C showsthe effect of anti PDL-1 treated tumor-derived cell mediated T cellproliferation.

FIGS. 56A and 56B show that cells isolated from tumors from Compound BBand anti-PDL-1 treated mice are less inhibitory of spleen T cellproliferation as compared to tumor derived cells from vehicle treatedmice.

FIG. 57 shows Compound BB reduces immune suppressive environment via anex vivo T-cell proliferation assay.

FIG. 58 shows LLC-bearing mice treated with Compound BB inhibits the exvivo whole blood stimulation of phospho-AKT in T cells.

FIG. 59 shows LLC-bearing mice treated with Compound BB inhibits the exvivo whole blood stimulation of phospho-AKT in monocytes.

FIG. 60 shows the effect of Compound BB on CXCL12- or anti-IgD-inducedpAKT in B cells.

FIG. 61 shows the single dose unbound plasma concentration of CompoundBB versus time profiles in male beagle dogs.

DETAILED DESCRIPTION

In one embodiment, provided are heterocyclyl compounds, andpharmaceutically acceptable forms thereof, including, but not limitedto, salts, hydrates, solvates, isomers, prodrugs, and isotopicallylabeled derivatives thereof.

In another embodiment, provided are methods of treating and/or managingvarious diseases and disorders, which comprises administering to apatient a therapeutically effective amount of a compound providedherein, or a pharmaceutically acceptable form (e.g., salts, hydrates,solvates, isomers, prodrugs, and isotopically labeled derivatives)thereof. Examples of diseases and disorders are described herein.

In another embodiment, provided are methods of preventing variousdiseases and disorders, which comprises administering to a patient inneed of such prevention a prophylactically effective amount of acompound provided herein, or a pharmaceutically acceptable form (e.g.,salts, hydrates, solvates, isomers, prodrugs, and isotopically labeledderivatives) thereof. Examples of diseases and disorders are describedherein.

In other embodiments, a compound provided herein, or a pharmaceuticallyacceptable form (e.g., salts, hydrates, solvates, isomers, prodrugs, andisotopically labeled derivatives) thereof, is administered incombination with another drug (“second active agent”) or treatment.Second active agents include small molecules and large molecules (e.g.,proteins and antibodies), examples of which are provided herein, as wellas stem cells. Other methods or therapies that can be used incombination with the administration of compounds provided hereininclude, but are not limited to, surgery, blood transfusions,immunotherapy, biological therapy, radiation therapy, and other non-drugbased therapies presently used to treat, prevent or manage variousdisorders described herein.

Also provided are pharmaceutical compositions (e.g., single unit dosageforms) that can be used in the methods provided herein. In oneembodiment, pharmaceutical compositions comprise a compound providedherein, or a pharmaceutically acceptable form (e.g., salts, hydrates,solvates, isomers, prodrugs, and isotopically labeled derivatives)thereof, and optionally one or more second active agents.

While specific embodiments have been discussed, the specification isillustrative only and not restrictive. Many variations of thisdisclosure will become apparent to those skilled in the art upon reviewof this specification.

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as is commonly understood by one of skill in theart to which this specification pertains.

As used in the specification and claims, the singular form “a”, “an” and“the” includes plural references unless the context clearly dictatesotherwise.

As used herein, and unless otherwise indicated, the term “about” or“approximately” means an acceptable error for a particular value asdetermined by one of ordinary skill in the art, which depends in part onhow the value is measured or determined. In certain embodiments, theterm “about” or “approximately” means within 1, 2, 3, or 4 standarddeviations. In certain embodiments, the term “about” or “approximately”means within 50%, 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%,0.5%, or 0.05% of a given value or range.

As used herein, “agent” or “biologically active agent” or “second activeagent” refers to a biological, pharmaceutical, or chemical compound orother moiety. Non-limiting examples include simple or complex organic orinorganic molecules, a peptide, a protein, an oligonucleotide, anantibody, an antibody derivative, an antibody fragment, a vitamin, avitamin derivative, a carbohydrate, a toxin, or a chemotherapeuticcompound, and metabolites thereof. Various compounds can be synthesized,for example, small molecules and oligomers (e.g., oligopeptides andoligonucleotides), and synthetic organic compounds based on various corestructures. In addition, various natural sources can provide compoundsfor screening, such as plant or animal extracts, and the like. A skilledartisan can readily recognize that there is no limit as to thestructural nature of the agents of this disclosure.

The term “agonist” as used herein refers to a compound or agent havingthe ability to initiate or enhance a biological function of a targetprotein or polypeptide, such as increasing the activity or expression ofthe target protein or polypeptide. Accordingly, the term “agonist” isdefined in the context of the biological role of the target protein orpolypeptide. While some agonists herein specifically interact with(e.g., bind to) the target, compounds and/or agents that initiate orenhance a biological activity of the target protein or polypeptide byinteracting with other members of the signal transduction pathway ofwhich the target polypeptide is a member are also specifically includedwithin this definition.

The terms “antagonist” and “inhibitor” are used interchangeably, andthey refer to a compound or agent having the ability to inhibit abiological function of a target protein or polypeptide, such as byinhibiting the activity or expression of the target protein orpolypeptide. Accordingly, the terms “antagonist” and “inhibitor” aredefined in the context of the biological role of the target protein orpolypeptide. While some antagonists herein specifically interact with(e.g., bind to) the target, compounds that inhibit a biological activityof the target protein or polypeptide by interacting with other membersof the signal transduction pathway of which the target protein orpolypeptide are also specifically included within this definition.Non-limiting examples of biological activity inhibited by an antagonistinclude those associated with the development, growth, or spread of atumor, or an undesired immune response as manifested in autoimmunedisease. The term “inhibition” or “inhibitor” as used in this contextincludes a reduction in a certain parameter, e.g., an activity, of agiven molecule, e.g., a PI3K isoform. For example, inhibition of anactivity, e.g., a PI3K activity, of at least 5%, 10%, 20%, 30%, 40% ormore is included by this term. Thus, inhibition need not be 100%.

An “anti-cancer agent”, “anti-tumor agent” or “chemotherapeutic agent”refers to any agent useful in the treatment of a neoplastic condition.One class of anti-cancer agents comprises chemotherapeutic agents.“Chemotherapy” means the administration of one or more chemotherapeuticdrugs and/or other agents to a cancer patient by various methods,including intravenous, oral, intramuscular, intraperitoneal,intravesical, subcutaneous, transdermal, or buccal administration, orinhalation, or in the form of a suppository.

The term “cell proliferation” refers to a phenomenon by which the cellnumber has changed as a result of division. This term also encompassescell growth by which the cell morphology has changed (e.g., increased insize) consistent with a proliferative signal.

The term “tumor” refers to any neoplastic cell growth and proliferation,whether malignant or benign, and any pre-cancerous and cancerous cellsand tissues. As used herein, the term “neoplastic” refers to any form ofdysregulated or unregulated cell growth, whether malignant or benign,resulting in abnormal tissue growth. Thus, “neoplastic cells” includemalignant and benign cells having dysregulated or unregulated cellgrowth.

The term “cancer” includes, but is not limited to, solid tumors andblood born tumors. The term “cancer” refers to disease of skin tissues,organs, blood, and vessels, including, but not limited to, cancers ofthe bladder, bone or blood, brain, breast, cervix, chest, colon,endometrium, esophagus, eye, head, kidney, liver, lymph nodes, lung,mouth, neck, ovaries, pancreas, prostate, rectum, stomach, testis,throat, and uterus.

Hematopoietic origin refers to involving cells generated duringhematopoiesis, a process by which cellular elements of blood, such aslymphocytes, leukocytes, platelets, erythrocytes and natural killercells are generated. Cancers of hematopoietic origin includes lymphomaand leukemia.

“Resistant” or “refractory” or “refractive” refers to when a cancer thathas a reduced responsiveness to a treatment, e.g., up to the point wherethe cancer does not respond to treatment. The cancer can be resistant atthe beginning of treatment, or it may become resistant during treatment.The cancer subject can have one or more mutations that cause it tobecome resistant to the treatment, or the subject may have developedsuch mutations during treatment. In one embodiment, the cancer orsubject has failed to respond to a given therapeutic treatment (e.g.,has failed to respond by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%,80%, 90%, or 100% to a given treatment). Failed treatment can bemeasured by, e.g., tumor volume or the length of time before tumorregrowth occurs.

By “hyperproliferative cancerous disease or disorder” is meant allneoplastic cell growth and proliferation, whether malignant or benign,including all transformed cells and tissues and all cancerous cells andtissues. Hyperproliferative diseases or disorders include, but are notlimited to, precancerous lesions, abnormal cell growth, benign tumors,malignant tumors, and “cancer.”

Combination therapy or “in combination with” refer to the use of morethan one compound or agent to treat a particular disorder or condition.For example, Compound 4 may be administered in combination with at leastone additional therapeutic agent. By “in combination with,” it is notintended to imply that the other therapy and Compound 4 must beadministered at the same time and/or formulated for delivery together,although these methods of delivery are within the scope of thisdisclosure. Compound 4 can be administered concurrently with, prior to(e.g., 5 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4hours, 6 hours, 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, 1week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, 12 weeks, or16 weeks before), or subsequent to (e.g., 5 minutes, 15 minutes, 30minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24hours, 48 hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks,5 weeks, 6 weeks, 8 weeks, 12 weeks, or 16 weeks after), one or moreother additional agents. In general, each therapeutic agent will beadministered at a dose and/or on a time schedule determined for thatparticular agent. The other therapeutic agent can be administered withCompound 4 herein in a single composition or separately in a differentcomposition. Higher combinations, e.g., triple therapy, are alsocontemplated herein.

As used herein, a “monotherapy” refers to the use of an agentindividually (e.g., as a single compound or agent), e.g., without asecond active ingredient to treat the same indication, e.g., cancer. Forexample, in this context, the term monotherapy includes the use ofeither the PI3K inhibitor or the second agent individually to treat thecancer.

The term “synergy” or “synergistic” encompasses a more than additiveeffect of a combination of two or more agents compared to theirindividual effects. In certain embodiments, synergy or synergisticeffect refers to an advantageous effect of using two or more agents incombination, e.g., in a pharmaceutical composition, or in a method oftreatment. In certain embodiments, one or more advantageous effects isachieved by using a PI3K inhibitor in combination with a secondtherapeutic agent (e.g., one or more second therapeutic agents) asdescribed herein.

In embodiments, the synergistic effect is that a lower dosage of one orboth of the agents is needed to achieve an effect. For example, thecombination can provide a selected effect, e.g., a therapeutic effect,when at least one of the agents is administered at a lower dosage thanthe dose of that agent that would be required to achieve the sametherapeutic effect when the agent is administered as a monotherapy. Incertain embodiments, the combination of a PI3K inhibitor (e.g., Compound4) and a second agent (as described herein) allows the PI3K inhibitor tobe administered at a lower dosage than would be required to achieve thesame therapeutic effect if the PI3K inhibitor were administered as amonotherapy.

In embodiments, a synergistic effect refers to the combination of a PI3Kinhibitor (e.g., Compound 4, or a pharmaceutically acceptable formthereof), and a second therapeutic agent (e.g., one or more additionaltherapeutic agent(s), or a pharmaceutically acceptable form thereof, asdescribed herein), results in a therapeutic effect greater than theadditive effect of the PI3K inhibitor and the second agent.

In embodiments, a synergistic effect means that combination index valueis less than a selected value, e.g., for a given effect, e.g., at aselected percentage (e.g., 50%) inhibition or growth inhibition, e.g.,as described herein in the Examples. In embodiments, a synergisticeffect means that the synergy score is 1 or more. In certainembodiments, the synergy score is greater than 1. In certainembodiments, the synergy score is greater than 3.

Combination index (CI) is a measure of potency shifting. The combinationindex is known in the art and is described, e.g., in Chou et al., AdvEnzyme Regul 1984; 22: 27-55 and in U.S. Patent Publication No.2013/0295102, the contents of which are incorporated herein byreference. A CI value of greater than 1 indicates antagonistic effect; aCI value of 1.0 is indicative of an additive effect; and a CI value ofless than 1 is indicative of a synergistic effect resulting from thecombination. The CI value can be determined at various percentages ofinhibition or growth inhibition.

The CI provides an estimate of the fraction of the original(monotherapy) doses of each of two drugs would be needed in combinationrelative to the single agent doses required to achieve a chosen effectlevel. For example, when the combination index has a value of 0.1, onlyabout one tenth of the total fractional amounts of the individual agents(expressed as a fraction of the amount of that agent when administeredas a monotherapy to achieve a chosen effect) are needed for thecombination to reach the same chosen effect level. For example, if adose of 100 mg/kg of drug A individually or a dose of 200 mg/kg of drugB individually is needed to achieve the chosen effect, and thecombination index is 0.1, then approximately 5 mg/kg of drug A and 10mg/kg of drug B would achieve the chosen effect (one twentieth of theoriginal doses of each of the single agents adds up to a total of onetenth). The doses of the single agents need not be reduced by the samefractional value so long as the sum of their fractional values adds upto the combination index; thus, in this example, a dose of approximately8 mg/kg of drug A and 4 mg/kg of drug B would also achieve the choseneffect (this is 0.08 times the original dose of drug A and 0.02 timesthe original dose of drug B; the sum of the fractional amounts(0.08+0.02) is equal to the combination index of 0.1.)

According to one embodiment, synergy score is a measure of thecombination effects in excess of Loewe additivity. In one example,synergy score is a scalar measure to characterize the strength ofsynergistic interaction. The Synergy score can be calculated as:Synergy Score=log f _(X) log f _(Y)Σ max(0,I _(data))(I _(data) −I_(Loewe))In this example, the fractional inhibition for each component agent andcombination point in the matrix is calculated relative to the median ofall vehicle-treated control wells. The example Synergy Score equationintegrates the experimentally-observed activity volume at each point inthe matrix in excess of a model surface numerically derived from theactivity of the component agents using the Loewe model for additivity.Additional terms in the Synergy Score equation (above) are used tonormalize for various dilution factors used for individual agents and toallow for comparison of synergy scores across an entire experiment. Theinclusion of positive inhibition gating or an I_(data) multiplierremoves noise near the zero effect level, and biases results forsynergistic interactions at that occur at high activity levels.According to other embodiments, a synergy score can be calculated basedon a curve fitting approach where the curvature of the synergy score isextrapolated by introducing a median value and origin value (e.g., adose zero value).

The synergy score measure can be used for the self-cross analysis.Synergy scores of self-crosses are expected to be additive by definitionand, therefore, maintain a synergy score of zero. However, while someself-cross synergy scores are near zero, many are greater suggestingthat experimental noise or non-optimal curve fitting of the single agentdose responses are contributing to the slight perturbations in thescore. This strategy is cell line-centric, focusing on self-crossbehavior in each cell line versus a global review of cell line panelactivity. Combinations where the synergy score is greater than the meanself-cross plus two standard deviations or three standard deviations canbe considered candidate synergies at 95% and 99% confidence levels,respectively. Additivity should maintain a synergy score of zero, andsynergy score of two or three standard deviations indicate synergism atstatistically significant levels of 95% and 99%.

Loewe Volume (Loewe Vol) can be used to assess the overall magnitude ofthe combination interaction in excess of the Loewe additivity model.Loewe Volume is particularly useful when distinguishing synergisticincreases in a phenotypic activity (positive Loewe Volume) versussynergistic antagonisms (negative Loewe Volume). When antagonisms areobserved, the Loewe Volume should be assessed to examine if there is anycorrelation between antagonism and a particular drug target-activity orcellular genotype. This model defines additivity as a non-synergisticcombination interaction where the combination dose matrix surface shouldbe indistinguishable from either drug crossed with itself. Thecalculation for Loewe additivity is:I _(Loewe) that satisfies(X/X _(I))+(Y/Y _(I))=1where X_(I) and Y_(I) are the single agent effective concentrations forthe observed combination effect I. For example, if 50% inhibition isachieved separately by 1 μM of drug A or 1 μM of drug B, a combinationof 0.5 μM of A and 0.5 μM of B should also inhibit by 50%.

The term “co-administration,” “administered in combination with,” andtheir grammatical equivalents, as used herein, encompass administrationof two or more agents to subject so that both agents and/or theirmetabolites are present in the subject at the same time.Co-administration includes simultaneous administration in separatecompositions, administration at different times in separatecompositions, or administration in a composition in which both agentsare present.

The term “effective amount” or “therapeutically effective amount” refersto that amount of a compound or pharmaceutical composition describedherein that is sufficient to effect the intended application including,but not limited to, disease treatment, as illustrated below. Thetherapeutically effective amount can vary depending upon the intendedapplication (in vitro or in vivo), or the subject and disease conditionbeing treated, e.g., the weight and age of the subject, the severity ofthe disease condition, the manner of administration and the like, whichcan readily be determined by one of ordinary skill in the art. The termalso applies to a dose that will induce a particular response in targetcells, e.g., reduction of platelet adhesion and/or cell migration. Thespecific dose will vary depending on, for example, the particularcompounds chosen, the dosing regimen to be followed, whether it isadministered in combination with other agents, timing of administration,the tissue to which it is administered, and the physical delivery systemin which it is carried.

As used herein, the terms “treatment”, “treating”, “palliating” and“ameliorating” are used interchangeably herein. These terms refer to anapproach for obtaining beneficial or desired results including, but notlimited to, therapeutic benefit. By therapeutic benefit is meanteradication or amelioration of the underlying disorder being treated.Also, a therapeutic benefit is achieved with the eradication oramelioration of one or more of the physiological symptoms associatedwith the underlying disorder such that an improvement is observed in thepatient, notwithstanding that the patient can still be afflicted withthe underlying disorder.

As used herein, the terms “prevention” and “preventing” are used hereinto refer to an approach for obtaining beneficial or desired resultsincluding, but not limited, to prophylactic benefit. For prophylacticbenefit, the pharmaceutical compositions can be administered to apatient at risk of developing a particular disease, or to a patientreporting one or more of the physiological symptoms of a disease, eventhough a diagnosis of this disease may not have been made.

A “therapeutic effect,” as that term is used herein, encompasses atherapeutic benefit and/or a prophylactic benefit as described above. Aprophylactic effect includes delaying or eliminating the appearance of adisease or condition, delaying or eliminating the onset of symptoms of adisease or condition, slowing, halting, or reversing the progression ofa disease or condition, or any combination thereof.

“Signal transduction” or “signaling pathway” is a process during whichstimulatory or inhibitory signals are transmitted into and within a cellto elicit an intracellular response. A “modulator” of a signaltransduction pathway refers to a compound which modulates the activityof one or more cellular proteins mapped to the same specific signaltransduction pathway. A modulator can augment (agonist) or suppress orinhibit (antagonist) the activity of a signaling molecule.

In certain embodiments, the signal transduction is mediated by one ormore phosphoinositide 3-kinases (PI3Ks). PI3Ks are members of aconserved family of lipid kinases that regulate numerous cell functions,including proliferation, differentiation, cell survival and metabolism.Several classes of PI3Ks exist in mammalian cells, including Class IAsubgroup (e.g., PI3K-α, β, δ), which are generally activated by receptortyrosine kinases (RTKs); Class IB (e.g., PI3K-γ), which is activated byG-protein coupled receptors (GPCRs), among others. PI3Ks exert theirbiological activities via a “PI3K-mediated signaling pathway” thatincludes several components that directly and/or indirectly transduce asignal triggered by a PI3K, including the generation of second messengerphophotidylinositol, 3,4,5-triphosphate (PIP3) at the plasma membrane,activation of heterotrimeric G protein signaling, and generation offurther second messengers such as cAMP, DAG, and IP3, all of which leadsto an extensive cascade of protein kinase activation (reviewed inVanhaesebroeck, B. et al. (2001) Annu Rev Biochem. 70:535-602). Incertain embodiments, the compounds disclosed herein inhibit a PI3 kinaseor PI3K) isoform, e.g., one, two, three or more of PI3K-α, β, δ or -γ.

In the context of biological molecules, to “decrease”, “suppress,”“ameliorate,” “reduce,” “inhibit,” or the like, includes decreasing alevel or an activity (e.g., one or more functions) of a given molecule.The level of a given molecule, e.g., mRNA or protein level, or theactivity can be measured in a sample, or using the assays described inthe Examples herein.

To “decrease,” “ameliorate,” “reduce,” “inhibit,” (or the like) adisorder or condition, or a symptom associated with a disorder orcondition includes reducing the severity and/or frequency of one or moresymptoms of the disorder or condition, or reducing or delaying the onsetof the disorder or condition and/or one or more symptoms of the disorderor condition. In some embodiments, the symptom is reduced by at leastabout 2%, at least about 5%, at least about 10%, at least about 15%, atleast about 20%, at least about 25%, at least about 30%, at least about40%, at least about 50%, at least about 60%, at least about 70%, atleast about 80%, at least about 90%, or at least about 95% relative to acontrol level.

The term “inhibition” or “inhibit” as used in this context includes areduction in a certain parameter, e.g., an activity, of a givenmolecule, e.g., a PI3K isoform. For example, inhibition of an activity,e.g., a PI3K activity, of at least 5%, 10%, 20%, 30%, 40% or more isincluded by this term. Thus, inhibition need not be 100%. In certainembodiments, a PI3K inhibitor as disclosed herein inhibits a PI3 kinaseof the gamma isoform (a “PI3K-γ isoform).

The term “selective inhibition” or “selectively inhibit” as applied to abiologically active agent refers to the agent's ability to selectivelyreduce the target signaling activity as compared to off-target signalingactivity, via direct or indirect interaction with the target. Forexample, a compound that selectively inhibits one isoform of PI3K overanother isoform of PI3K has an activity of at least greater than about1× against a first isoform relative to the compound's activity againstthe second isoform (e.g., at least about 2×, 3×, 5×, 10×, 20×, 50×,100×, 200×, 500×, or 1000×). In certain embodiments, these terms referto (1) a compound of described herein that selectively inhibits thegamma isoform over the alpha, beta, or delta isoform; or (2) a compounddescribed herein that selectively inhibits the delta isoform over thealpha or beta isoform. By way of non-limiting example, the ratio ofselectivity can be greater than a factor of about 1, greater than afactor of about 2, greater than a factor of about 3, greater than afactor of about 5, greater than a factor of about 10, greater than afactor of about 50, greater than a factor of about 100, greater than afactor of about 200, greater than a factor of about 400, greater than afactor of about 600, greater than a factor of about 800, greater than afactor of about 1000, greater than a factor of about 1500, greater thana factor of about 2000, greater than a factor of about 5000, greaterthan a factor of about 10,000, or greater than a factor of about 20,000,where selectivity can be measured by ratio of IC50 values, which in turncan be measured by, e.g., in vitro or in vivo assays such as thosedescribed in Examples described herein. In one embodiment, theselectivity of a first PI3K isoform over a second PI3K isoform ismeasured by the ratio of the IC50 value against the second PI3K isoformto the IC50 value against the first PI3K gamma isoform. For example, adelta/gamma selectivity ratio of a compound can be measured by the ratioof the compound's inhibitory activity against the delta isoform in termsof IC50 or the like to the compound's inhibitory activity against thegamma isoform in terms of IC50 or the like. If the delta/gammaselectivity ratio is larger than 1, the compound selectively inhibitsthe gamma isoform over the delta isoform. In certain embodiments, thePI3K gamma isoform IC50 activity of a compound of provided herein can beless than about 1000 nM, less than about 500 nM, less than about 400 nM,less than about 300 nM, less than about 200 nM, less than about 100 nM,less than about 75 nM, less than about 50 nM, less than about 25 nM,less than about 20 nM, less than about 15 nM, less than about 10 nM,less than about 5 nM, or less than about 1 nM. In certain embodiments,the PI3K delta isoform IC50 activity of a compound provided herein canbe less than about 1000 nM, less than about 500 nM, less than about 400nM, less than about 300 nM, less than about 200 nM, less than about 100nM, less than about 75 nM, less than about 50 nM, less than about 25 nM,less than about 20 nM, less than about 15 nM, less than about 10 nM,less than about 5 nM, or less than about 1 nM.

In certain embodiments, a PI3K-γ inhibitor selectively inhibits thegamma isoform over the alpha, beta, or delta isoform (also referred toherein as a “PI3K-γ-selective inhibitor.” In one embodiment, the PI3K-γinhibitor selectively inhibits the gamma isoform over the alpha or betaisoform. In one embodiment, the PI3K-γ inhibitor selectively inhibitsthe gamma isoform over the alpha, beta, and delta isoforms. In oneembodiment, the PI3K-γ inhibitor selectively inhibits the gamma isoformover the alpha and beta isoforms. In one embodiment, the PI3K-γinhibitor selectively inhibits the gamma isoform over the alpha and betaisoforms, but not the delta isoform. By way of non-limiting example, theratio of selectivity can be greater than a factor of about 10, greaterthan a factor of about 50, greater than a factor of about 100, greaterthan a factor of about 200, greater than a factor of about 400, greaterthan a factor of about 600, greater than a factor of about 800, greaterthan a factor of about 1000, greater than a factor of about 1500,greater than a factor of about 2000, greater than a factor of about5000, greater than a factor of about 10,000, or greater than a factor ofabout 20,000, where selectivity can be measured by ratio of IC50 values,among other means. In one embodiment, the selectivity of PI3K gammaisoform over another PI3K isoform is measured by the ratio of the IC50value against the other PI3K isoform to the IC50 value against PI3Kgamma isoform. In certain embodiments, the PI3 kinase gamma isoform IC50activity of a compound as disclosed herein can be less than about 1000nM, less than about 100 nM, less than about 10 nM, or less than about 1nM. For example, a compound that selectively inhibits one isoform ofPI3K over another isoform of PI3K has an activity of at least 2× againsta first isoform relative to the compound's activity against the secondisoform (e.g., at least about 3×, 5×, 10×, 20×, 50×, 100×, 200×, 500×,or 1000×).

In certain embodiments, a PI3K-γ selective inhibitor is used oradministered to a subject at a lower dose (e.g., by about 10%, by about20%, by about 30%, by about 40%, by about 50%, by about 60%, by about70%, or by about 80%) as compared to treatment with a PI3K-γnon-selective or less selective PI3K-γ inhibitor (e.g., a PI3Kpaninhibitors, e.g., inhibiting PI3K-α, β, δ, and γ).

“Radiation therapy” means exposing a patient to radiation emitters suchas, but not limited to, alpha-particle emitting radionuclides (e.g.,actinium and thorium radionuclides), low linear energy transfer (LET)radiation emitters (e.g., beta emitters), conversion electron emitters(e.g., strontium-89 and samarium-153-EDTMP), or high-energy radiation,including without limitation x-rays, gamma rays, and neutrons. Radiationtherapy can be performed using routine methods and compositions known tothe practitioner.

As used herein, a “reference value” refers to a value to which a givenvalue can be compared. In some embodiments, the reference value refersto a control (e.g., an untreated control, e.g., an untreated orplacebo-treated subject or an untreated sample); the course of diseasewithout treatment; a healthy subject or an average of healthy subjects;a subject at a different time interval, e.g., prior to, during, or afterthe treatment).

“Subject” to which administration is contemplated includes, but is notlimited to, humans (e.g., a male or female of any age group, e.g., apediatric subject (e.g., infant, child, adolescent) or adult subject(e.g., young adult, middle-aged adult or senior adult)) and/or otherprimates (e.g., cynomolgus monkeys, rhesus monkeys); mammals, includingcommercially relevant mammals such as cattle, pigs, horses, sheep,goats, cats, and/or dogs; and/or birds, including commercially relevantbirds such as chickens, ducks, geese, quail, and/or turkeys.

The term “in vivo” refers to an event that takes place in a subject'sbody.

The term “in vitro” refers to an event that takes places outside of asubject's body. For example, an in vitro assay encompasses any assayconducted outside of a subject. In vitro assays encompass cell-basedassays in which cells, alive or dead, are employed. In vitro assays alsoencompass a cell-free assay in which no intact cells are employed.

Chemical Definitions

As used herein, “pharmaceutically acceptable esters” include, but arenot limited to, alkyl, alkenyl, alkynyl, aryl, aralkyl, and cycloalkylesters of acidic groups, including, but not limited to, carboxylicacids, phosphoric acids, phosphinic acids, sulfonic acids, sulfinicacids, and boronic acids.

As used herein, “pharmaceutically acceptable enol ethers” include, butare not limited to, derivatives of formula —C═C(OR) where R can beselected from alkyl, alkenyl, alkynyl, aryl, aralkyl, and cycloalkyl.Pharmaceutically acceptable enol esters include, but are not limited to,derivatives of formula —C═C(OC(O)R) where R can be selected fromhydrogen, alkyl, alkenyl, alkynyl, aryl, aralkyl, and cycloalkyl.

As used herein, a “pharmaceutically acceptable form” of a disclosedcompound includes, but is not limited to, pharmaceutically acceptablesalts, hydrates, solvates, isomers, prodrugs, and isotopically labeledderivatives of disclosed compounds. In one embodiment, a“pharmaceutically acceptable form” includes, but is not limited to,pharmaceutically acceptable salts, isomers, prodrugs and isotopicallylabeled derivatives of disclosed compounds.

In certain embodiments, the pharmaceutically acceptable form is apharmaceutically acceptable salt. As used herein, the term“pharmaceutically acceptable salt” refers to those salts which are,within the scope of sound medical judgment, suitable for use in contactwith the tissues of subjects without undue toxicity, irritation,allergic response and the like, and are commensurate with a reasonablebenefit/risk ratio. Pharmaceutically acceptable salts are well known inthe art. For example, Berge et al. describes pharmaceutically acceptablesalts in detail in J. Pharmaceutical Sciences (1977) 66:1-19.Pharmaceutically acceptable salts of the compounds provided hereininclude those derived from suitable inorganic and organic acids andbases. Examples of pharmaceutically acceptable, nontoxic acid additionsalts are salts of an amino group formed with inorganic acids such ashydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid andperchloric acid or with organic acids such as acetic acid, oxalic acid,maleic acid, tartaric acid, citric acid, succinic acid or malonic acidor by using other methods used in the art such as ion exchange. Otherpharmaceutically acceptable salts include adipate, alginate, ascorbate,aspartate, benzenesulfonate, besylate, benzoate, bisulfate, borate,butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate,digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate,glucoheptonate, glycerophosphate, gluconate, hemisulfate, heptanoate,hexanoate, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate,lactate, laurate, lauryl sulfate, malate, maleate, malonate,methanesulfonate, 2-naphthalenesulfonate, naphthalene-m,n-bissulfonates,nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate,persulfate, 3-phenylpropionate, phosphate, picrate, pivalate,propionate, stearate, succinate, sulfate, tartrate, thiocyanate,p-toluenesulfonate, undecanoate, valerate salts, and the like. In someembodiments, organic acids from which salts can be derived include, forexample, acetic acid, propionic acid, glycolic acid, pyruvic acid,oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid,tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid,methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid,salicylic acid, naphthalene-m,n-bissulfonic acids and the like.

Pharmaceutically acceptable salts derived from appropriate bases includealkali metal, alkaline earth metal, ammonium and N⁺(C₁₋₄alkyl)₄ salts.Representative alkali or alkaline earth metal salts include sodium,lithium, potassium, calcium, magnesium, iron, zinc, copper, manganese,aluminum, and the like. Further pharmaceutically acceptable saltsinclude, when appropriate, nontoxic ammonium, quaternary ammonium, andamine cations formed using counterions such as halide, hydroxide,carboxylate, sulfate, phosphate, nitrate, lower alkyl sulfonate, andaryl sulfonate. Organic bases from which salts can be derived include,for example, primary, secondary, and tertiary amines, substituted aminesincluding naturally occurring substituted amines, cyclic amines, basicion exchange resins, and the like, such as isopropylamine,trimethylamine, diethylamine, triethylamine, tripropylamine, andethanolamine. In some embodiments, the pharmaceutically acceptable baseaddition salt is chosen from ammonium, potassium, sodium, calcium, andmagnesium salts.

In certain embodiments, the pharmaceutically acceptable form is asolvate (e.g., a hydrate). As used herein, the term “solvate” refers tocompounds that further include a stoichiometric or non-stoichiometricamount of solvent bound by non-covalent intermolecular forces. Thesolvate can be of a disclosed compound or a pharmaceutically acceptablesalt thereof. Where the solvent is water, the solvate is a “hydrate”.Pharmaceutically acceptable solvates and hydrates are complexes that,for example, can include 1 to about 100, or 1 to about 10, or one toabout 2, about 3 or about 4, solvent or water molecules. It will beunderstood that the term “compound” as used herein encompasses thecompound and solvates of the compound, as well as mixtures thereof.

In certain embodiments, the pharmaceutically acceptable form is aprodrug. As used herein, the term “prodrug” refers to compounds that aretransformed in vivo to yield a disclosed compound or a pharmaceuticallyacceptable form of the compound. A prodrug can be inactive whenadministered to a subject, but is converted in vivo to an activecompound, for example, by hydrolysis (e.g., hydrolysis in blood). Incertain cases, a prodrug has improved physical and/or deliveryproperties over the parent compound. Prodrugs are typically designed toenhance pharmaceutically and/or pharmacokinetically based propertiesassociated with the parent compound. The prodrug compound often offersadvantages of solubility, tissue compatibility or delayed release in amammalian organism (see, e.g., Bundgard, H., Design of Prodrugs (1985),pp. 7-9, 21-24 (Elsevier, Amsterdam). A discussion of prodrugs isprovided in Higuchi, T., et al., “Pro-drugs as Novel Delivery Systems,”A.C.S. Symposium Series, Vol. 14, and in Bioreversible Carriers in DrugDesign, ed. Edward B. Roche, American Pharmaceutical Association andPergamon Press, 1987, both of which are incorporated in full byreference herein. Exemplary advantages of a prodrug can include, but arenot limited to, its physical properties, such as enhanced watersolubility for parenteral administration at physiological pH compared tothe parent compound, or it enhances absorption from the digestive tract,or it can enhance drug stability for long-term storage.

The term “prodrug” is also meant to include any covalently bondedcarriers, which release the active compound in vivo when such prodrug isadministered to a subject. Prodrugs of an active compound, as describedherein, can be prepared by modifying functional groups present in theactive compound in such a way that the modifications are cleaved, eitherin routine manipulation or in vivo, to the parent active compound.Prodrugs include compounds wherein a hydroxy, amino or mercapto group isbonded to any group that, when the prodrug of the active compound isadministered to a subject, cleaves to form a free hydroxy, free amino orfree mercapto group, respectively. Examples of prodrugs include, but arenot limited to, acetate, formate and benzoate derivatives of an alcoholor acetamide, formamide and benzamide derivatives of an amine functionalgroup in the active compound and the like. Other examples of prodrugsinclude compounds that comprise —NO, —NO₂, —ONO, or —ONO₂ moieties.Prodrugs can typically be prepared using well-known methods, such asthose described in Burger's Medicinal Chemistry and Drug Discovery,172-178, 949-982 (Manfred E. Wolff ed., 5th ed., 1995), and Design ofProdrugs (H. Bundgaard ed., Elsevier, New York, 1985).

For example, if a disclosed compound or a pharmaceutically acceptableform of the compound contains a carboxylic acid functional group, aprodrug can comprise a pharmaceutically acceptable ester formed by thereplacement of the hydrogen atom of the acid group with a group such as(C₁-C₈)alkyl, (C₂-C₁₂)alkanoyloxymethyl, 1-(alkanoyloxy)ethyl havingfrom 4 to 9 carbon atoms, 1-methyl-1-(alkanoyloxy)-ethyl having from 5to 10 carbon atoms, alkoxycarbonyloxymethyl having from 3 to 6 carbonatoms, 1-(alkoxycarbonyloxy)ethyl having from 4 to 7 carbon atoms,1-methyl-1-(alkoxycarbonyloxy)ethyl having from 5 to 8 carbon atoms,N-(alkoxycarbonyl)aminomethyl having from 3 to 9 carbon atoms,1-(N-(alkoxycarbonyl)amino)ethyl having from 4 to 10 carbon atoms,3-phthalidyl, 4-crotonolactonyl, gamma-butyrolacton-4-yl,di-N,N—(C₁-C₂)alkylamino(C₂-C₃)alkyl (such as β-dimethylaminoethyl),carbamoyl-(C₁-C₂)alkyl, N,N-di(C₁-C₂)alkylcarbamoyl-(C₁-C₂)alkyl andpiperidino-, pyrrolidino- or morpholino(C₂-C₃)alkyl.

Similarly, if a disclosed compound or a pharmaceutically acceptable formof the compound contains an alcohol functional group, a prodrug can beformed by the replacement of the hydrogen atom of the alcohol group witha group such as (C₁-C₆)alkanoyloxymethyl, 1-((C₁-C₆)alkanoyloxy)ethyl,1-methyl-1-((C₁-C₆)alkanoyloxy)ethyl (C₁-C₆)alkoxycarbonyloxymethyl,N—(C₁-C₆)alkoxycarbonylaminomethyl, succinoyl, (C₁-C₆)alkanoyl,α-amino(C₁-C₄)alkanoyl, arylacyl and α-aminoacyl, orα-aminoacyl-α-aminoacyl, where each α-aminoacyl group is independentlyselected from naturally occurring L-amino acids, P(O)(OH)₂,—P(O)(O(C₁-C₆)alkyl)₂, and glycosyl (the radical resulting from theremoval of a hydroxyl group of the hemiacetal form of a carbohydrate).

If a disclosed compound or a pharmaceutically acceptable form of thecompound incorporates an amine functional group, a prodrug can be formedby the replacement of a hydrogen atom in the amine group with a groupsuch as R-carbonyl, RO-carbonyl, NRR′-carbonyl where R and R′ are eachindependently (C₁-C₁₀)alkyl, (C₃-C₇)cycloalkyl, benzyl, a naturalα-aminoacyl or natural α-aminoacyl-natural α-aminoacyl, —C(OH)C(O)OY¹wherein Y¹ is H, (C₁-C₆)alkyl or benzyl, —C(OY²)Y³ wherein Y² is (C₁-C₄)alkyl and Y³ is (C₁-C₆)alkyl, carboxy(C₁-C₆)alkyl, amino(C₁-C₄)alkyl ormono-N— or di-N,N—(C₁-C₆)alkylaminoalkyl, —C(Y⁴)Y⁵ wherein Y⁴ is H ormethyl and Y⁵ is mono-N— or di-N,N—(C₁-C₆)alkylamino, morpholino,piperidin-1-yl or pyrrolidin-1-yl.

In certain embodiments, the pharmaceutically acceptable form is anisomer. “Isomers” are different compounds that have the same molecularformula. “Atropisomers” are stereoisomers from hindered rotation aboutsingle bonds and can be resolved or isolated by methods known to thoseskilled in the art. For example, certain B substituents of a compound ofFormula (I) provided herein with ortho or meta substituted phenyl mayform atropisomers, where they may be separated and isolated.

“Stereoisomers” are isomers that differ only in the way the atoms arearranged in space. As used herein, the term “isomer” includes any andall geometric isomers and stereoisomers. For example, “isomers” includegeometric double bond cis- and trans-isomers, also termed E- andZ-isomers; R- and S-enantiomers; diastereomers, (d)-isomers and(l)-isomers, racemic mixtures thereof; and other mixtures thereof, asfalling within the scope of this disclosure.

In certain embodiments, the symbol

denotes a bond that can be a single or double as described herein.

In certain embodiments, provided herein are various geometric isomersand mixtures thereof resulting from the arrangement of substituentsaround a carbon-carbon double bond or arrangement of substituents arounda carbocyclic ring. Substituents around a carbon-carbon double bond aredesignated as being in the “Z” or “E” configuration wherein the terms“Z” and “E” are used in accordance with IUPAC standards. Unlessotherwise specified, structures depicting double bonds encompass boththe “E” and “Z” isomers.

Substituents around a carbon-carbon double bond alternatively can bereferred to as “cis” or “trans,” where “cis” represents substituents onthe same side of the double bond and “trans” represents substituents onopposite sides of the double bond. The arrangement of substituentsaround a carbocyclic ring can also be designated as “cis” or “trans.”The term “cis” represents substituents on the same side of the plane ofthe ring, and the term “trans” represents substituents on opposite sidesof the plane of the ring. Mixtures of compounds wherein the substituentsare disposed on both the same and opposite sides of the plane of thering are designated “cis/trans.”

“Enantiomers” are a pair of stereoisomers that are non-superimposablemirror images of each other. A mixture of a pair of enantiomers in anyproportion can be known as a “racemic” mixture. The term “(±)” is usedto designate a racemic mixture where appropriate. “Diastereoisomers” arestereoisomers that have at least two asymmetric atoms, but which are notmirror-images of each other. The absolute stereochemistry can bespecified according to the Cahn-Ingold-Prelog R-S system. When acompound is an enantiomer, the stereochemistry at each chiral carbon canbe specified by either R or S. Resolved compounds whose absoluteconfiguration is unknown can be designated (+) or (−) depending on thedirection (dextro- or levorotatory) which they rotate plane polarizedlight at the wavelength of the sodium D line. Certain of the compoundsdescribed herein contain one or more asymmetric centers and can thusgive rise to enantiomers, diastereomers, and other stereoisomeric formsthat can be defined, in terms of absolute stereochemistry at eachasymmetric atom, as (R)- or (S)-. The present chemical entities,pharmaceutical compositions and methods are meant to include all suchpossible isomers, including racemic mixtures, optically substantiallypure forms and intermediate mixtures. Optically active (R)- and(S)-isomers can be prepared, for example, using chiral synthons orchiral reagents, or resolved using conventional techniques.

The “enantiomeric excess” or “% enantiomeric excess” of a compositioncan be calculated using the equation shown below. In the example shownbelow, a composition contains 90% of one enantiomer, e.g., an Senantiomer, and 10% of the other enantiomer, e.g., an R enantiomer.ee=(90−10)/100=80%.

Thus, a composition containing 90% of one enantiomer and 10% of theother enantiomer is said to have an enantiomeric excess of 80%. Somecompositions described herein contain an enantiomeric excess of at leastabout 1%, about 5%, about 10%, about 20%, about 30%, about 40%, about50%, about 75%, about 90%, about 95%, or about 99% of the S enantiomer.In other words, the compositions contain an enantiomeric excess of the Senantiomer over the R enantiomer. In other embodiments, somecompositions described herein contain an enantiomeric excess of at leastabout 1%, about 5%, about 10%, about 20%, about 30%, about 40%, about50%, about 75%, about 90%, about 95%, or about 99% of the R enantiomer.In other words, the compositions contain an enantiomeric excess of the Renantiomer over the S enantiomer.

For instance, an isomer/enantiomer can, in some embodiments, be providedsubstantially free of the corresponding enantiomer, and can also bereferred to as “optically enriched,” “enantiomerically enriched,”“enantiomerically pure” and “non-racemic,” as used interchangeablyherein. These terms refer to compositions in which the amount of oneenantiomer is greater than the amount of that one enantiomer in acontrol mixture of the racemic composition (e.g., greater than 1:1 byweight). For example, an enantiomerically enriched preparation of the Senantiomer, means a preparation of the compound having greater thanabout 50% by weight of the S enantiomer relative to the total weight ofthe preparation (e.g., total weight of S and R isomers). such as atleast about 75% by weight, further such as at least about 80% by weight.In some embodiments, the enrichment can be much greater than about 80%by weight, providing a “substantially enantiomerically enriched,”“substantially enantiomerically pure” or a “substantially non-racemic”preparation, which refers to preparations of compositions which have atleast about 85% by weight of one enantiomer relative to the total weightof the preparation, such as at least about 90% by weight, and furthersuch as at least about 95% by weight. In certain embodiments, thecompound provided herein is made up of at least about 90% by weight ofone enantiomer. In other embodiments, the compound is made up of atleast about 95%, about 98%, or about 99% by weight of one enantiomer.

In some embodiments, the compound is a racemic mixture of (S)- and(R)-isomers. In other embodiments, provided herein is a mixture ofcompounds wherein individual compounds of the mixture existpredominately in an (S)- or (R)-isomeric configuration. For example, insome embodiments, the compound mixture has an (S)-enantiomeric excess ofgreater than about 10%, greater than about 20%, greater than about 30%,greater than about 40%, greater than about 50%, greater than about 55%,greater than about 60%, greater than about 65%, greater than about 70%,greater than about 75%, greater than about 80%, greater than about 85%,greater than about 90%, greater than about 95%, greater than about 96%,greater than about 97%, greater than about 98%, or greater than about99%. In some embodiments, the compound mixture has an (S)-enantiomericexcess of about 55%, about 60%, about 65%, about 70%, about 75%, about80%, about 85%, about 90%, about 95%, about 96%, about 97%, about 98%,about 99%, or about 99.5%, or more. In some embodiments, the compoundmixture has an (S)-enantiomeric excess of about 55% to about 99.5%,about 60% to about 99.5%, about 65% to about 99.5%, about 70% to about99.5%, about 75% to about 99.5%, about 80% to about 99.5%, about 85% toabout 99.5%, about 90% to about 99.5%, about 95% to about 99.5%, about96% to about 99.5%, about 97% to about 99.5%, about 98% to about 99.5%,or about 99% to about 99.5%, or more than about 99.5%.

In other embodiments, the compound mixture has an (R)-enantiomericexcess of greater than about 10%, greater than about 20%, greater thanabout 30%, greater than about 40%, greater than about 50%, greater thanabout 55%, greater than about 60%, greater than about 65%, greater thanabout 70%, greater than about 75%, greater than about 80%, greater thanabout 85%, greater than about 90%, greater than about 95%, greater thanabout 96%, greater than about 97%, greater than about 98%, or greaterthan about 99%. In some embodiments, the compound mixture has an(R)-enantiomeric excess of about 55%, about 60%, about 65%, about 70%,about 75%, about 80%, about 85%, about 90%, about 95%, about 96%, about97%, about 98%, about 99%, or about 99.5%, or more. In some embodiments,the compound mixture has an (R)-enantiomeric excess of about 55% toabout 99.5%, about 60% to about 99.5%, about 65% to about 99.5%, about70% to about 99.5%, about 75% to about 99.5%, about 80% to about 99.5%,about 85% to about 99.5%, about 90% to about 99.5%, about 95% to about99.5%, about 96% to about 99.5%, about 97% to about 99.5%, about 98% toabout 99.5%, or about 99% to about 99.5%, or more than about 99.5%.

In other embodiments, the compound mixture contains identical chemicalentities except for their stereochemical orientations, namely (S)- or(R)-isomers. For example, if a compound disclosed herein has —CH(R)—unit, and R is not hydrogen, then the —CH(R)— is in an (S)- or(R)-stereochemical orientation for each of the identical chemicalentities (i.e., (S)- or (R)-stereoisomers). In some embodiments, themixture of identical chemical entities (i.e., mixture of stereoisomers)is a racemic mixture of (S)- and (R)-isomers. In another embodiment, themixture of the identical chemical entities (i.e., mixture ofstereoisomers) contains predominately (S)-isomer or predominately(R)-isomer. For example, in some embodiments, the (S)-isomer in themixture of identical chemical entities (i.e., mixture of stereoisomers)is present at about 55%, about 60%, about 65%, about 70%, about 75%,about 80%, about 85%, about 90%, about 95%, about 96%, about 9′7%, about98%, about 99%, or about 99.5% by weight, or more, relative to the totalweight of the mixture of (S)- and (R)-isomers. In some embodiments, the(S)-isomer in the mixture of identical chemical entities (i.e., mixtureof stereoisomers) is present at an (S)-enantiomeric excess of about 10%to about 99.5%, about 20% to about 99.5%, about 30% to about 99.5%,about 40% to about 99.5%, about 50% to about 99.5%, about 55% to about99.5%, about 60% to about 99.5%, about 65% to about 99.5%, about 70% toabout 99.5%, about 75% to about 99.5%, about 80% to about 99.5%, about85% to about 99.5%, about 90% to about 99.5%, about 95% to about 99.5%,about 96% to about 99.5%, about 97% to about 99.5%, about 98% to about99.5%, or about 99% to about 99.5%, or more than about 99.5%.

In other embodiments, the (R)-isomer in the mixture of identicalchemical entities (i.e., mixture of stereoisomers) is present at about55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%,about 90%, about 95%, about 96%, about 97%, about 98%, about 99%, orabout 99.5% by weight, or more, relative to the total weight of themixture of (S)- and (R)-isomers. In some embodiments, the (R)-isomers inthe mixture of identical chemical entities (i.e., mixture ofstereoisomers) is present at an (R)-enantiomeric excess of about 10% toabout 99.5%, about 20% to about 99.5%, about 30% to about 99.5%, about40% to about 99.5%, about 50% to about 99.5%, about 55% to about 99.5%,about 60% to about 99.5%, about 65% to about 99.5%, about 70% to about99.5%, about 75% to about 99.5%, about 80% to about 99.5%, about 85% toabout 99.5%, about 90% to about 99.5%, about 95% to about 99.5%, about96% to about 99.5%, about 97% to about 99.5%, about 98% to about 99.5%,or about 99% to about 99.5%, or more than about 99.5%.

Enantiomers can be isolated from racemic mixtures by any method known tothose skilled in the art, including chiral high pressure liquidchromatography (HPLC), the formation and crystallization of chiralsalts, or prepared by asymmetric syntheses. See, for example,Enantiomers, Racemates and Resolutions (Jacques, Ed., WileyInterscience, New York, 1981); Wilen et al., Tetrahedron 33:2725 (1977);Stereochemistry of Carbon Compounds (E. L. Eliel, Ed., McGraw-Hill, N Y,1962); and Tables of Resolving Agents and Optical Resolutions p. 268 (E.L. Eliel, Ed., Univ. of Notre Dame Press, Notre Dame, Ind. 1972).

In certain embodiments, the pharmaceutically acceptable form is atautomer. As used herein, the term “tautomer” is a type of isomer thatincludes two or more interconvertable compounds resulting from at leastone formal migration of a hydrogen atom and at least one change invalency (e.g., a single bond to a double bond, a triple bond to a doublebond, or a triple bond to a single bond, or vice versa).“Tautomerization” includes prototropic or proton-shift tautomerization,which is considered a subset of acid-base chemistry. “Prototropictautomerization” or “proton-shift tautomerization” involves themigration of a proton accompanied by changes in bond order. The exactratio of the tautomers depends on several factors, includingtemperature, solvent, and pH. Where tautomerization is possible (e.g.,in solution), a chemical equilibrium of tautomers can be reached.Tautomerizations (i.e., the reaction providing a tautomeric pair) can becatalyzed by acid or base, or can occur without the action or presenceof an external agent. Exemplary tautomerizations include, but are notlimited to, keto-enol; amide-imide; lactam-lactim; enamine-imine; andenamine-(a different) enamine tautomerizations. A specific example ofketo-enol tautomerization is the interconversion of pentane-2,4-dioneand 4-hydroxypent-3-en-2-one tautomers. Another example oftautomerization is phenol-keto tautomerization. A specific example ofphenol-keto tautomerization is the interconversion of pyridin-4-ol andpyridin-4(1H)-one tautomers.

Unless otherwise stated, structures depicted herein are also meant toinclude compounds which differ only in the presence of one or moreisotopically enriched atoms. For example, compounds having the presentstructures except for the replacement or enrichment of a hydrogen bydeuterium or tritium at one or more atoms in the molecule, or thereplacement or enrichment of a carbon by ¹³C or ¹⁴C at one or more atomsin the molecule, are within the scope of this disclosure. In oneembodiment, provided herein are isotopically labeled compounds havingone or more hydrogen atoms replaced by or enriched by deuterium. In oneembodiment, provided herein are isotopically labeled compounds havingone or more hydrogen atoms replaced by or enriched by tritium. In oneembodiment, provided herein are isotopically labeled compounds havingone or more carbon atoms replaced or enriched by ¹³C. In one embodiment,provided herein are isotopically labeled compounds having one or morecarbon atoms replaced or enriched by ¹⁴C.

The disclosure also embraces isotopically labeled compounds which areidentical to those recited herein, except that one or more atoms arereplaced by an atom having an atomic mass or mass number different fromthe atomic mass or mass number usually found in nature. Examples ofisotopes that can be incorporated into disclosed compounds includeisotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur,fluorine, and chlorine, such as, e.g., ²H, ³H, ¹³C, ¹⁴C, ¹⁵N, ¹⁸O, ¹⁷O,³¹P, ³²P, ³⁵S, ¹⁸F, and ³⁶Cl, respectively. Certain isotopically-labeleddisclosed compounds (e.g., those labeled with ³H and/or ¹⁴C) are usefulin compound and/or substrate tissue distribution assays. Tritiated(i.e., ³H) and carbon-14 (i.e., ¹⁴C) isotopes can allow for ease ofpreparation and detectability. Further, substitution with heavierisotopes such as deuterium (i.e., ²H) can afford certain therapeuticadvantages resulting from greater metabolic stability (e.g., increasedin vivo half-life or reduced dosage requirements). Isotopically labeleddisclosed compounds can generally be prepared by substituting anisotopically labeled reagent for a non-isotopically labeled reagent. Insome embodiments, provided herein are compounds that can also containunnatural proportions of atomic isotopes at one or more of atoms thatconstitute such compounds. All isotopic variations of the compounds asdisclosed herein, whether radioactive or not, are encompassed within thescope of the present disclosure.

“Pharmaceutically acceptable carrier” or “pharmaceutically acceptableexcipient” includes any and all solvents, dispersion media, coatings,antibacterial and antifungal agents, isotonic and absorption delayingagents and the like. The use of such media and agents forpharmaceutically active substances is well known in the art. Exceptinsofar as any conventional media or agent is incompatible with theactive ingredient, its use in the therapeutic compositions as disclosedherein is contemplated. Supplementary active ingredients can also beincorporated into the pharmaceutical compositions.

Definitions of specific functional groups and chemical terms aredescribed in more detail below. The chemical elements are identified inaccordance with the Periodic Table of the Elements, CAS version,Handbook of Chemistry and Physics, 75th ed., inside cover, and specificfunctional groups are generally defined as described therein.Additionally, general principles of organic chemistry, as well asspecific functional moieties and reactivity, are described in OrganicChemistry, Thomas Sorrell, University Science Books, Sausalito, 1999;Smith and March March's Advanced Organic Chemistry, 5th ed., John Wiley& Sons, Inc., New York, 2001; Larock, Comprehensive OrganicTransformations, VCH Publishers, Inc., New York, 1989; and Carruthers,Some Modern Methods of Organic Synthesis, 3rd ed., Cambridge UniversityPress, Cambridge, 1987.

When a range of values is listed, it is intended to encompass each valueand sub-range within the range. For example “C₁₋₆ alkyl” is intended toencompass, C₁, C₂, C₃, C₄, C₅, C₆, C₁₋₆, C₁₋₅, C₁₋₄, C₁₋₃, C₁₋₂, C₂₋₆,C₂₋₅, C₂₋₄, C₂₋₃, C₃₋₆, C₃₋₅, C₃₋₄, C₄₋₆, C₄₋₅, and C₅₋₆ alkyl.

“Alkyl” refers to a straight or branched hydrocarbon chain radicalconsisting solely of carbon and hydrogen atoms, containing nounsaturation, having, in some embodiments, from one to ten carbon atoms(e.g., C₁-C₁₀ alkyl). Linear or straight alkyl refers to an alkyl withno branching, e.g., methyl, ethyl, n-propyl. Whenever it appears herein,a numerical range such as “1 to 10” refers to each integer in the givenrange; e.g., “1 to 10 carbon atoms” means that the alkyl group canconsist of 1 carbon atom, 2 carbon atoms, 3 carbon atoms, 4 carbonatoms, etc., up to and including 10 carbon atoms, although the presentdefinition also covers the occurrence of the term “alkyl” where nonumerical range is designated. In some embodiments, an alkyl is a C₁-C₆alkyl group. In some embodiments, alkyl groups have 1 to 10, 1 to 6, 1to 4, or 1 to 3 carbon atoms. Representative saturated straight chainalkyls include, but are not limited to, -methyl, -ethyl, -n-propyl,-n-butyl, -n-pentyl, and -n-hexyl; while saturated branched alkylsinclude, but are not limited to, -isopropyl, -sec-butyl, -isobutyl,-tert-butyl, -isopentyl, 2-methylbutyl, 3-methylbutyl, 2-methylpentyl,3-methylpentyl, 4-methylpentyl, 2-methylhexyl, 3-methylhexyl,4-methylhexyl, 5-methylhexyl, 2,3-dimethylbutyl, and the like. The alkylis attached to the parent molecule by a single bond. Unless statedotherwise in the specification, an alkyl group is optionally substitutedby one or more of substituents which independently include: acyl, alkyl,alkenyl, alkynyl, alkoxy, alkylaryl, cycloalkyl, aralkyl, aryl, aryloxy,amino, amido, amidino, imino, azide, carbonate, carbamate, carbonyl,heteroalkyl, heteroaryl, heteroarylalkyl, heterocycloalkyl, hydroxy,cyano, halo, haloalkoxy, haloalkyl, ester, ether, mercapto, thio,alkylthio, arylthio, thiocarbonyl, nitro, oxo, phosphate, phosphonate,phosphinate, silyl, sulfinyl, sulfonyl, sulfonamidyl, sulfoxyl,sulfonate, urea, —Si(R^(a))₃, —OR^(a), —SR^(a), —OC(O)—R^(a),—N(R^(a))₂, —C(O)R^(a), —C(O)OR^(a), —OC(O)N(R^(a))₂, —C(O)N(R^(a))₂,—N(R^(a))C(O)OR^(a), —N(R^(a))C(O)R^(a), —N(R^(a))C(O)N(R^(a))₂,—N(R^(a))C(NR^(a))N(R^(a))₂, —N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2),—S(O)_(t)OR^(a) (where t is 1 or 2), —S(O)_(t)N(R^(a))₂ (where t is 1 or2), or —O—P(═O)(OR^(a))₂, where each R^(a) is independently hydrogen,alkyl, haloalkyl, carbocyclyl, carbocyclylalkyl, aryl, aralkyl,heterocycloalkyl, heterocycloalkylalkyl, heteroaryl or heteroarylalkyl,and each of these moieties can be optionally substituted as definedherein.

“Perhaloalkyl” refers to an alkyl group in which all of the hydrogenatoms have been replaced with a halogen selected from fluoro, chloro,bromo, and iodo. In some embodiments, all of the hydrogen atoms are eachreplaced with fluoro. In some embodiments, all of the hydrogen atoms areeach replaced with chloro. Examples of perhaloalkyl groups include —CF₃,—CF₂CF₃, —CF₂CF₂CF₃, —CCl₃, —CFCl₂, —CF₂Cl and the like. “Haloalkyl”refers to an alkyl group in which one or more of the hydrogen atoms havebeen replaced with a halogen independently selected from fluoro, chloro,bromo, and iodo.

“Alkyl-cycloalkyl” refers to an -(alkyl)cycloalkyl radical where alkyland cycloalkyl are as disclosed herein and which are optionallysubstituted by one or more of the substituents described as suitablesubstituents for alkyl and cycloalkyl respectively. The“alkyl-cycloalkyl” is bonded to the parent molecular structure throughthe alkyl group. The terms “alkenyl-cycloalkyl” and “alkynyl-cycloalkyl”mirror the above description of “alkyl-cycloalkyl” wherein the term“alkyl” is replaced with “alkenyl” or “alkynyl” respectively, and“alkenyl” or “alkynyl” are as described herein.

“Alkylaryl” refers to an -(alkyl)aryl radical where aryl and alkyl areas disclosed herein and which are optionally substituted by one or moreof the substituents described as suitable substituents for aryl andalkyl respectively. The “alkylaryl” is bonded to the parent molecularstructure through the alkyl group. The terms “-(alkenyl)aryl” and“-(alkynyl)aryl” mirror the above description of “-(alkyl)aryl” whereinthe term “alkyl” is replaced with “alkenyl” or “alkynyl” respectively,and “alkenyl” or “alkynyl” are as described herein.

“Alkyl-heteroaryl” refers to an -(alkyl)heteroaryl radical whereheteroaryl and alkyl are as disclosed herein and which are optionallysubstituted by one or more of the substituents described as suitablesubstituents for heteroaryl and alkyl respectively. The“alkyl-heteroaryl” is bonded to the parent molecular structure throughthe alkyl group. The terms “-(alkenyl)heteroaryl” and“-(alkynyl)heteroaryl” mirror the above description of“-(alkyl)heteroaryl” wherein the term “alkyl” is replaced with “alkenyl”or “alkynyl” respectively, and “alkenyl” or “alkynyl” are as describedherein.

“Alkyl-heterocyclyl” refers to an -(alkyl)heterocyclyl radical wherealkyl and heterocyclyl are as disclosed herein and which are optionallysubstituted by one or more of the substituents described as suitablesubstituents for heterocyclyl and alkyl respectively. The“alkyl-heterocyclyl” is bonded to the parent molecular structure throughthe alkyl group. The terms “-(alkenyl)heterocyclyl” and“-(alkynyl)heterocyclyl” mirror the above description of“-(alkyl)heterocyclyl” wherein the term “alkyl” is replaced with“alkenyl” or “alkynyl” respectively, and “alkenyl” or “alkynyl” are asdescribed herein.

“Alkenyl” refers to a straight or branched hydrocarbon chain radicalgroup consisting solely of carbon and hydrogen atoms, containing atleast one double bond, and in some embodiments, having from two to tencarbon atoms (i.e., C₂-C₁₀ alkenyl). Whenever it appears herein, anumerical range such as “2 to 10” refers to each integer in the givenrange; e.g., “2 to 10 carbon atoms” means that the alkenyl group canconsist of 2 carbon atoms, 3 carbon atoms, 4 carbon atoms, etc., up toand including 10 carbon atoms. In certain embodiments, an alkenylcomprises two to eight carbon atoms. In other embodiments, an alkenylcomprises two to five carbon atoms (e.g., C₂-C₅ alkenyl). The alkenyl isattached to the parent molecular structure by a single bond, forexample, ethenyl (i.e., vinyl), prop-1-enyl (i.e., allyl), but-1-enyl,pent-1-enyl, penta-1,4-dienyl, and the like. The one or morecarbon-carbon double bonds can be internal (such as in 2-butenyl) orterminal (such as in 1-butenyl). Examples of C₂₋₄ alkenyl groups includeethenyl (C₂), 1-propenyl (C₃), 2-propenyl (C₃), 1-butenyl (C₄),2-butenyl (C₄), butadienyl (C₄) and the like. Examples of C₂₋₆ alkenylgroups include the aforementioned C₂₋₄ alkenyl groups as well aspentenyl (C₅), pentadienyl (C₅), hexenyl (C₆), and the like. Additionalexamples of alkenyl include heptenyl (C₇), octenyl (C₈), octatrienyl(C₈), and the like. Unless stated otherwise in the specification, analkenyl group is optionally substituted by one or more substituentswhich independently include: acyl, alkyl, alkenyl, alkynyl, alkoxy,alkylaryl, cycloalkyl, aralkyl, aryl, aryloxy, amino, amido, amidino,imino, azide, carbonate, carbamate, carbonyl, heteroalkyl, heteroaryl,heteroarylalkyl, heterocycloalkyl, hydroxy, cyano, halo, haloalkoxy,haloalkyl, ester, ether, mercapto, thio, alkylthio, arylthio,thiocarbonyl, nitro, oxo, phosphate, phosphonate, phosphinate, silyl,sulfinyl, sulfonyl, sulfonamidyl, sulfoxyl, sulfonate, urea,—Si(R^(a))₃, —OR^(a), —SR^(a), —OC(O)—R^(a), —N(R^(a))₂, —C(O)R^(a),—C(O)OR^(a), —OC(O)N(R^(a))₂, —C(O)N(R^(a))₂, —N(R^(a))C(O)OR^(a),—N(R^(a))C(O)R^(a), —N(R^(a))C(O)N(R^(a))₂, —N(R^(a))C(NR^(a))N(R^(a))₂,—N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2), —S(O)_(t)OR^(a) (where t is1 or 2), —S(O)_(t)N(R^(a))₂ (where t is 1 or 2), or —O—P(═O)(OR^(a))₂,where each R^(a) is independently hydrogen, alkyl, haloalkyl,carbocyclyl, carbocyclylalkyl, aryl, aralkyl, heterocycloalkyl,heterocycloalkylalkyl, heteroaryl or heteroarylalkyl, and each of thesemoieties can be optionally substituted as defined herein.

“Alkynyl” refers to a straight or branched hydrocarbon chain radicalgroup consisting solely of carbon and hydrogen atoms, containing atleast one triple bond, having, in some embodiments, from two to tencarbon atoms (i.e., C₂-C₁₀ alkynyl). Whenever it appears herein, anumerical range such as “2 to 10” refers to each integer in the givenrange; e.g., “2 to 10 carbon atoms” means that the alkynyl group canconsist of 2 carbon atoms, 3 carbon atoms, 4 carbon atoms, etc., up toand including 10 carbon atoms. In certain embodiments, an alkynylcomprises two to eight carbon atoms. In other embodiments, an alkynylhas two to five carbon atoms (e.g., C₂-C₅ alkynyl). The alkynyl isattached to the parent molecular structure by a single bond, forexample, ethynyl, propynyl, butynyl, pentynyl, hexynyl, and the like.Unless stated otherwise in the specification, an alkynyl group isoptionally substituted by one or more substituents which independentlyinclude: acyl, alkyl, alkenyl, alkynyl, alkoxy, alkylaryl, cycloalkyl,aralkyl, aryl, aryloxy, amino, amido, amidino, imino, azide, carbonate,carbamate, carbonyl, heteroalkyl, heteroaryl, heteroarylalkyl,heterocycloalkyl, hydroxy, cyano, halo, haloalkoxy, haloalkyl, ester,ether, mercapto, thio, alkylthio, arylthio, thiocarbonyl, nitro, oxo,phosphate, phosphonate, phosphinate, silyl, sulfinyl, sulfonyl,sulfonamidyl, sulfoxyl, sulfonate, urea, —Si(R^(a))₃, —OR^(a), —SR^(a),—OC(O)—R^(a), —N(R^(a))₂, —C(O)R^(a), —C(O)OR^(a), —OC(O)N(R^(a))₂,—C(O)N(R^(a))₂, —N(R^(a))C(O)OR^(a), —N(R^(a))C(O)R^(a),—N(R^(a))C(O)N(R^(a))₂, —N(R^(a))C(NR^(a))N(R^(a))₂,—N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2), —S(O)_(t)OR^(a) (where t is1 or 2), —S(O)_(t)N(R^(a))₂ (where t is 1 or 2), or —O—P(═O)(OR^(a))₂,where each R^(a) is independently hydrogen, alkyl, haloalkyl,carbocyclyl, carbocyclylalkyl, aryl, aralkyl, heterocycloalkyl,heterocycloalkylalkyl, heteroaryl or heteroarylalkyl, and each of thesemoieties can be optionally substituted as defined herein.

The term “alkoxy” refers to the group —O-alkyl (in some embodiments,including from 1 to 10 carbon atoms), of a straight, branched, cyclicconfiguration and combinations thereof, attached to the parent molecularstructure through an oxygen. Examples include methoxy, ethoxy, propoxy,isopropoxy, cyclopropyloxy, cyclohexyloxy, and the like. “Lower alkoxy”refers to alkoxy groups containing one to six carbons. In someembodiments, C₁-C₄ alkoxy is an alkoxy group which encompasses bothstraight and branched chain alkyls of from 1 to 4 carbon atoms. Unlessstated otherwise in the specification, an alkoxy group is optionallysubstituted by one or more substituents which independently include:acyl, alkyl, alkenyl, alkynyl, alkoxy, alkylaryl, cycloalkyl, aralkyl,aryl, aryloxy, amino, amido, amidino, imino, azide, carbonate,carbamate, carbonyl, heteroalkyl, heteroaryl, heteroarylalkyl,heterocycloalkyl, hydroxy, cyano, halo, haloalkoxy, haloalkyl, ester,ether, mercapto, thio, alkylthio, arylthio, thiocarbonyl, nitro, oxo,phosphate, phosphonate, phosphinate, silyl, sulfinyl, sulfonyl,sulfonamidyl, sulfoxyl, sulfonate, urea, —Si(R^(a))₃, —OR^(a), —SR^(a),—OC(O)—R^(a), —N(R^(a))₂, —C(O)R^(a), —C(O)OR^(a), —OC(O)N(R^(a))₂,—C(O)N(R^(a))₂, —N(R^(a))C(O)OR^(a), —N(R^(a))C(O)R^(a),—N(R^(a))C(O)N(R^(a))₂, —N(R^(a))C(NR^(a))N(R^(a))₂,—N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2), —S(O)_(t)OR^(a) (where t is1 or 2), —S(O)₁N(R^(a))₂ (where t is 1 or 2), or —O—P(═O)(R^(a))₂, whereeach R^(a) is independently hydrogen, alkyl, haloalkyl, carbocyclyl,carbocyclylalkyl, aryl, aralkyl, heterocycloalkyl,heterocycloalkylalkyl, heteroaryl or heteroarylalkyl, and each of thesemoieties can be optionally substituted as defined herein. The terms“alkenoxy” and “alkynoxy” mirror the above description of “alkoxy”wherein the prefix “alk” is replaced with “alken” or “alkyn”respectively, and the parent “alkenyl” or “alkynyl” terms are asdescribed herein.

The term “alkoxycarbonyl” refers to a group of the formula(alkoxy)(C═O)— attached to the parent molecular structure through thecarbonyl carbon (in some embodiments, having from 1 to 10 carbon atoms).Thus a C₁-C₆ alkoxycarbonyl group comprises an alkoxy group having from1 to 6 carbon atoms attached through its oxygen to a carbonyl linker.The C₁-C₆ designation does not include the carbonyl carbon in the atomcount. “Lower alkoxycarbonyl” refers to an alkoxycarbonyl group whereinthe alkyl portion of the alkoxy group is a lower alkyl group. In someembodiments, C₁-C₄ alkoxycarbonyl comprises an alkoxy group whichencompasses both straight and branched chain alkoxy groups of from 1 to4 carbon atoms. Unless stated otherwise in the specification, analkoxycarbonyl group is optionally substituted by one or moresubstituents which independently include: acyl, alkyl, alkenyl, alkynyl,alkoxy, alkylaryl, cycloalkyl, aralkyl, aryl, aryloxy, amino, amido,amidino, imino, azide, carbonate, carbamate, carbonyl, heteroalkyl,heteroaryl, heteroarylalkyl, heterocycloalkyl, hydroxy, cyano, halo,haloalkoxy, haloalkyl, ester, ether, mercapto, thio, alkylthio,arylthio, thiocarbonyl, nitro, oxo, phosphate, phosphonate, phosphinate,silyl, sulfinyl, sulfonyl, sulfonamidyl, sulfoxyl, sulfonate, urea,—Si(R^(a))₃, —OR^(a), —SR^(a), —OC(O)—R^(a), —N(R^(a))₂, —C(O)R^(a),—C(O)OR^(a), —OC(O)N(R^(a))₂, —C(O)N(R^(a))₂, —N(R^(a))C(O)OR^(a),—N(R^(a))C(O)R^(a), —N(R^(a))C(O)N(R^(a))₂,—N(R^(a))C(NR^(a))ON(R^(a))₂, —N(R^(a))S(O)_(t)R^(a) (where t is 1 or2), —S(O)_(t)OR^(a) (where t is 1 or 2), —S(O)_(t)N(R^(a))₂ (where t is1 or 2), or —O—P(═O)(OR^(a))₂, where each R^(a) is independentlyhydrogen, alkyl, haloalkyl, carbocyclyl, carbocyclylalkyl, aryl,aralkyl, heterocycloalkyl, heterocycloalkylalkyl, heteroaryl, orheteroarylalkyl, and each of these moieties can be optionallysubstituted as defined herein. The terms “alkenoxycarbonyl” and“alkynoxycarbonyl” mirror the above description of “alkoxycarbonyl”wherein the prefix “alk” is replaced with “alken” or “alkyn”respectively, and the parent “alkenyl” or “alkynyl” terms are asdescribed herein.

“Acyl” refers to R—C(O)— groups such as, but not limited to, H,(alkyl)-C(O)—, (alkenyl)-C(O)—, (alkynyl)-C(O)—, (aryl)-C(O)—,(cycloalkyl)-C(O)—, (heteroaryl)-C(O)—, (heteroalkyl)-C(O)—, and(heterocycloalkyl)-C(O)—, wherein the group is attached to the parentmolecular structure through the carbonyl functionality. In someembodiments, provided herein is a C₁-C₁₀ acyl radical which refers tothe total number of chain or ring atoms of the, for example, alkyl,alkenyl, alkynyl, aryl, cyclohexyl, heteroaryl or heterocycloalkylportion plus the carbonyl carbon of acyl. For example, a C4-acyl hasthree other ring or chain atoms plus carbonyl. If the R radical isheteroaryl or heterocycloalkyl, the hetero ring or chain atomscontribute to the total number of chain or ring atoms. Unless statedotherwise in the specification, the “R” of an acyloxy group can beoptionally substituted by one or more substituents which independentlyinclude: acyl, alkyl, alkenyl, alkynyl, alkoxy, alkylaryl, cycloalkyl,aralkyl, aryl, aryloxy, amino, amido, amidino, imino, azide, carbonate,carbamate, carbonyl, heteroalkyl, heteroaryl, heteroarylalkyl,heterocycloalkyl, hydroxy, cyano, halo, haloalkoxy, haloalkyl, ester,ether, mercapto, thio, alkylthio, arylthio, thiocarbonyl, nitro, oxo,phosphate, phosphonate, phosphinate, silyl, sulfinyl, sulfonyl,sulfonamidyl, sulfoxyl, sulfonate, urea, —Si(R^(a))₃, —OR^(a), —SR^(a),—OC(O)—R^(a), —N(R^(a))₂, —C(O)R^(a), —C(O)OR^(a), —OC(O)N(R^(a))₂,—C(O)N(R^(a))₂, —N(R^(a))C(O)OR^(a), —N(R^(a))C(O)R^(a),—N(R^(a))C(O)N(R^(a))₂, —N(R^(a))C(NR^(a))N(R^(a))₂,—N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2), —S(O)_(t)OR^(a) (where t is1 or 2), —S(O)_(t)N(R^(a))₂ (where t is 1 or 2), or —O—P(═O)(OR^(a))₂,where each R^(a) is independently hydrogen, alkyl, haloalkyl,carbocyclyl, carbocyclylalkyl, aryl, aralkyl, heterocycloalkyl,heterocycloalkylalkyl, heteroaryl, or heteroarylalkyl, and each of thesemoieties can be optionally substituted as defined herein.

“Acyloxy” refers to a R(C═O)O— radical wherein “R” can be H, alkyl,alkenyl, alkynyl, heteroalkyl, heteroalkenyl, heteroalkynyl, aryl,cyclohexyl, heteroaryl, or heterocycloalkyl, which are as describedherein. The acyloxy group is attached to the parent molecular structurethrough the oxygen functionality. In some embodiments, an acyloxy groupis a C₁-C₄ acyloxy radical which refers to the total number of chain orring atoms of the alkyl, alkenyl, alkynyl, aryl, cyclohexyl, heteroarylor heterocycloalkyl portion of the acyloxy group plus the carbonylcarbon of acyl, e.g., a C₄-acyloxy has three other ring or chain atomsplus carbonyl. If the R radical is heteroaryl or heterocycloalkyl, thehetero ring or chain atoms contribute to the total number of chain orring atoms. Unless stated otherwise in the specification, the “R” of anacyloxy group is optionally substituted by one or more substituentswhich independently include: acyl, alkyl, alkenyl, alkynyl, alkoxy,alkylaryl, cycloalkyl, aralkyl, aryl, aryloxy, amino, amido, amidino,imino, azide, carbonate, carbamate, carbonyl, heteroalkyl, heteroaryl,heteroarylalkyl, heterocycloalkyl, hydroxy, cyano, halo, haloalkoxy,haloalkyl, ester, ether, mercapto, thio, alkylthio, arylthio,thiocarbonyl, nitro, oxo, phosphate, phosphonate, phosphinate, silyl,sulfinyl, sulfonyl, sulfonamidyl, sulfoxyl, sulfonate, urea,—Si(R^(a))₃, —OR^(a), —OC(O)—R^(a), —N(R^(a))₂, —C(O)R^(a), —C(O)OR^(a),—OC(O)N(R^(a))₂, —C(O)N(R^(a))₂, —N(R^(a))C(O)OR^(a),—N(R^(a))C(O)R^(a), —N(R^(a))C(O)N(R^(a))₂, —N(R^(a))C(NR^(a))N(R^(a))₂,—N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2), —S(O)_(t)OR^(a) (where t is1 or 2), —S(O)_(t)N(R^(a))₂ (where t is 1 or 2), or —O—P(═O)(OR^(a))₂,where each R^(a) is independently hydrogen, alkyl, haloalkyl,carbocyclyl, carbocyclylalkyl, aryl, aralkyl, heterocycloalkyl,heterocycloalkylalkyl, heteroaryl, or heteroarylalkyl and each of thesemoieties can be optionally substituted as defined herein.

“Amino” or “amine” refers to a —N(R^(b))₂, —N(R^(b))R^(b)—, or—R^(b)N(R^(b))R^(b)— radical group, where each R^(b) is independentlyselected from hydrogen, alkyl, alkenyl, alkynyl, haloalkyl, heteroalkyl(bonded through a chain carbon), cycloalkyl, cycloalkylalkyl, aryl,aralkyl, heterocycloalkyl (bonded through a ring carbon),heterocycloalkylalkyl, heteroaryl (bonded through a ring carbon), andheteroarylalkyl, unless stated otherwise in the specification, each ofwhich moiety can itself be optionally substituted as described herein.When a —N(R^(b))₂ group has two R^(b) other than hydrogen, they can becombined with the nitrogen atom to form a 3-, 4-, 5-, 6-, 7-, or8-membered ring. For example, —N(R^(b))₂ is meant to include, but not belimited to, 1-pyrrolidinyl and 4-morpholinyl. Unless stated otherwise inthe specification, an amino group is optionally substituted by one ormore substituents which independently include: acyl, alkyl, alkenyl,alkynyl, alkoxy, alkylaryl, cycloalkyl, aralkyl, aryl, aryloxy, amino,amido, amidino, imino, azide, carbonate, carbamate, carbonyl,heteroalkyl, heteroaryl, heteroarylalkyl, heterocycloalkyl, hydroxy,cyano, halo, haloalkoxy, haloalkyl, ester, ether, mercapto, thio,alkylthio, arylthio, thiocarbonyl, nitro, oxo, phosphate, phosphonate,phosphinate, silyl, sulfinyl, sulfonyl, sulfonamidyl, sulfoxyl,sulfonate, urea, —Si(R^(a))₃, —OR^(a), —SR^(a), —OC(O)—R^(a),—N(R^(a))₂, —C(O)R^(a), —C(O)OR^(a), —OC(O)N(R^(a))₂, —C(O)N(R^(a))₂,—N(R^(a))C(O)OR^(a), —N(R^(a))C(O)R^(a), —N(R^(a))C(O)N(R^(a))₂,—N(R^(a))C(NR^(a))N(R^(a))₂, —N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2),—S(O)_(t)OR^(a) (where t is 1 or 2), —S(O)_(t)N(R^(a))₂ (where t is 1 or2), or —O—P(═O)(OR^(a))₂, where each R^(a) is independently hydrogen,alkyl, haloalkyl, carbocyclyl, carbocyclylalkyl, aryl, aralkyl,heterocycloalkyl, heterocycloalkylalkyl, heteroaryl, or heteroarylalkyl,and each of these moieties can be optionally substituted as definedherein.

The terms “amine” and “amino” can also refer to N-oxides of the groups—N⁺(H)(R^(a))O⁻, and —N⁺(R^(a))(R^(a))O⁻, where R^(a) is as describedabove, where the N-oxide is bonded to the parent molecular structurethrough the N atom. N-oxides can be prepared by treatment of thecorresponding amino group with, for example, hydrogen peroxide orm-chloroperoxybenzoic acid. The person skilled in the art is familiarwith reaction conditions for carrying out the N-oxidation.

“Amide” or “amido” refers to a chemical moiety with formula—C(O)N(R^(b))₂ or —NR^(b)C(O)R^(b), where R^(b) is independentlyselected from hydrogen, alkyl, alkenyl, alkynyl, haloalkyl, heteroalkyl(bonded through a chain carbon), cycloalkyl, cycloalkylalkyl, aryl,aralkyl, heterocycloalkyl (bonded through a ring carbon),heterocycloalkylalkyl, heteroaryl (bonded through a ring carbon), andheteroarylalkyl, unless stated otherwise in the specification, each ofwhich moiety can itself be optionally substituted as described herein.In some embodiments, an amido or amide radical is a C₁-C₄ amido or amideradical, which includes the amide carbonyl in the total number ofcarbons in the radical. When a —C(O)N(R^(b))₂ has two R^(b) other thanhydrogen, they can be combined with the nitrogen atom to form a 3-, 4-,5-, 6-, 7-, or 8-membered ring. For example, N(R^(b))₂ portion of a—C(O)N(R^(b))₂ radical is meant to include, but not be limited to,1-pyrrolidinyl and 4-morpholinyl. Unless stated otherwise in thespecification, an amido R^(b) group is optionally substituted by one ormore substituents which independently include: acyl, alkyl, alkenyl,alkynyl, alkoxy, alkylaryl, cycloalkyl, aralkyl, aryl, aryloxy, amino,amido, amidino, imino, azide, carbonate, carbamate, carbonyl,heteroalkyl, heteroaryl, heteroarylalkyl, heterocycloalkyl, hydroxy,cyano, halo, haloalkoxy, haloalkyl, ester, ether, mercapto, thio,alkylthio, arylthio, thiocarbonyl, nitro, oxo, phosphate, phosphonate,phosphinate, silyl, sulfinyl, sulfonyl, sulfonamidyl, sulfoxyl,sulfonate, urea, —Si(R^(a))₃, —OR^(a), —SR^(a), —OC(O)—R^(a),—N(R^(a))₂, —C(O)R^(a), —C(O)OR^(a), —OC(O)N(R^(a))₂, —C(O)N(R^(a))₂,—N(R^(a))C(O)OR^(a), —N(R^(a))C(O)R^(a), —N(R^(a))C(O)N(R^(a))₂,—N(R^(a))C(NR^(a))N(R^(a))₂, —N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2),—S(O)_(t)OR^(a) (where t is 1 or 2), —S(O)_(t)N(R^(a))₂ (where t is 1 or2), or —O—P(═O)(OR^(a))₂, where each R^(a) is independently hydrogen,alkyl, haloalkyl, carbocyclyl, carbocyclylalkyl, aryl, aralkyl,heterocycloalkyl, heterocycloalkylalkyl, heteroaryl, or heteroarylalkyl,and each of these moieties can be optionally substituted as definedherein.

The term “amide” or “amido” is inclusive of an amino acid or a peptidemolecule. Any amine, hydroxy, or carboxyl side chain on the compoundsdescribed herein can be transformed into an amide group. The proceduresand specific groups to make such amides are known to those of skill inthe art and can readily be found in reference sources such as Greene andWuts, Protective Groups in Organic Synthesis, 4th Ed., John Wiley &Sons, New York, N.Y., 2006, which is incorporated herein by reference inits entirety.

“Amidino” refers to the —C(═NR^(b))N(R^(b))₂,—N(R^(b))—C(═NR^(b))—R^(b), and —N(R^(b))—C(═NR^(b))— radicals, whereeach R^(b) is independently selected from hydrogen, alkyl, alkenyl,alkynyl, haloalkyl, heteroalkyl (bonded through a chain carbon),cycloalkyl, cycloalkylalkyl, aryl, aralkyl, heterocycloalkyl (bondedthrough a ring carbon), heterocycloalkylalkyl, heteroaryl (bondedthrough a ring carbon), and heteroarylalkyl, unless stated otherwise inthe specification, each of which moiety can itself be optionallysubstituted as described herein.

“Aryl” refers to a radical with six to fourteen ring atoms (e.g., C₆-C₁₄or C₆-C₁₀ aryl) which has at least one carbocyclic ring having aconjugated pi electron system which is aromatic (e.g., having 6, 10, or14 π electrons shared in a cyclic array) (e.g., phenyl, fluorenyl, andnaphthyl). In one embodiment, bivalent radicals formed from substitutedbenzene derivatives and having the free valences at ring atoms are namedas substituted phenylene radicals. In other embodiments, bivalentradicals derived from univalent monocyclic or polycyclic hydrocarbonradicals whose names end in “-yl” by removal of one hydrogen atom fromthe carbon atom with the free valence are named by adding “-idene” tothe name of the corresponding univalent radical, e.g., a naphthyl groupwith two points of attachment is termed naphthylidene. Whenever itappears herein, a numerical range such as “6 to 10 aryl” refers to eachinteger in the given range; e.g., “6 to 10 ring atoms” means that thearyl group can consist of 6 ring atoms, 7 ring atoms, etc., up to andincluding 10 ring atoms. The term includes monocyclic or fused-ringpolycyclic (i.e., rings which share adjacent pairs of ring atoms)groups. Unless stated otherwise in the specification, an aryl moiety canbe optionally substituted by one or more substituents whichindependently include: acyl, alkyl, alkenyl, alkynyl, alkoxy, alkylaryl,cycloalkyl, aralkyl, aryl, aryloxy, amino, amido, amidino, imino, azide,carbonate, carbamate, carbonyl, heteroalkyl, heteroaryl,heteroarylalkyl, heterocycloalkyl, hydroxy, cyano, halo, haloalkoxy,haloalkyl, ester, ether, mercapto, thio, alkylthio, arylthio,thiocarbonyl, nitro, oxo, phosphate, phosphonate, phosphinate, silyl,sulfinyl, sulfonyl, sulfonamidyl, sulfoxyl, sulfonate, urea,—Si(R^(a))₃, —OR^(a), —SR^(a), —OC(O)—R^(a), —N(R^(a))₂, —C(O)R^(a),—C(O)OR^(a), —OC(O)N(R^(a))₂, —C(O)N(R^(a))₂, —N(R^(a))C(O)OR^(a),—N(R^(a))C(O)R^(a), —N(R^(a))C(O)N(R^(a))₂, —N(R^(a))C(NR^(a))N(R^(a))₂,—N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2), —S(O)_(t)OR^(a) (where t is1 or 2), —S(O)_(t)N(R^(a))₂ (where t is 1 or 2), or —O—P(═O)(OR^(a))₂,where each R^(a) is independently hydrogen, alkyl, haloalkyl,carbocyclyl, carbocyclylalkyl, aryl, aralkyl, heterocycloalkyl,heterocycloalkylalkyl, heteroaryl, or heteroarylalkyl, and each of thesemoieties can be optionally substituted as defined herein. In oneembodiment, unless stated otherwise, “aryl” also includes ring systemswherein the aryl ring, as defined above, is fused with one or morecycloalkyl or heterocyclyl groups wherein the point of attachment to theparent molecular structure is on the aryl ring.

“Aralkyl” or “arylalkyl” refers to an (aryl)alkyl-radical where aryl andalkyl are as disclosed herein and which are optionally substituted byone or more of the substituents described as suitable substituents foraryl and alkyl respectively. The “aralkyl” or “arylalkyl” is bonded tothe parent molecular structure through the alkyl group. The terms“aralkenyl/arylalkenyl” and “aralkynyl/arylalkynyl” mirror the abovedescription of “aralkyl/arylalkyl” wherein the “alkyl” is replaced with“alkenyl” or “alkynyl” respectively, and the “alkenyl” or “alkynyl”terms are as described herein.

“Azide” refers to a —N₃ radical.

“Carbamate” refers to any of the following radicals: —O—(C═O)—N(R^(b))—,—O—(C═O)—N(R^(b))₂, —N(R^(b))—(C═O)—O—, and —N(R^(b))—(C═O)—OR^(b),wherein each R^(b) is independently selected from H, alkyl, alkenyl,alkynyl, haloalkyl, heteroalkyl (bonded through a chain carbon),cycloalkyl, cycloalkylalkyl, aryl, aralkyl, heterocycloalkyl (bondedthrough a ring carbon), heterocycloalkylalkyl, heteroaryl (bondedthrough a ring carbon), and heteroarylalkyl, unless stated otherwise inthe specification, each of which moiety can itself be optionallysubstituted as described herein.

“Carbonate” refers to a —O—(C═O)—O— or —O—(C═O)—OR radical, where R canbe hydrogen, alkyl, alkenyl, alkynyl, heteroalkyl, heteroalkenyl,heteroalkynyl, aryl, cyclohexyl, heteroaryl, or heterocycloalkyl, whichare as described herein.

“Carbonyl” refers to a —(C═O)— radical.

“Carboxaldehyde” refers to a —(C═O)H radical.

“Carboxyl” refers to a —(C═O)OH radical.

“Cyano” refers to a —CN radical.

“Cycloalkyl,” or alternatively, “carbocyclyl,” refers to a monocyclic orpolycyclic radical that contains only carbon and hydrogen, and can besaturated or partially unsaturated. Partially unsaturated cycloalkylgroups can be termed “cycloalkenyl” if the carbocycle contains at leastone double bond, or “cycloalkynyl” if the carbocycle contains at leastone triple bond. Cycloalkyl groups include groups having from 3 to 10ring atoms (e.g., C₃-C₁₀ cycloalkyl). Whenever it appears herein, anumerical range such as “3 to 10” refers to each integer in the givenrange; e.g., “3 to 10 carbon atoms” means that the cycloalkyl group canconsist of 3 carbon atoms, 4 carbon atoms, 5 carbon atoms, etc., up toand including 10 carbon atoms. The term “cycloalkyl” also includesbridged and spiro-fused cyclic structures containing no heteroatoms. Theterm also includes monocyclic or fused-ring polycyclic (i.e., ringswhich share adjacent pairs of ring atoms) groups. In some embodiments,it is a C₃-C₈ cycloalkyl radical. In some embodiments, it is a C₃-C₅cycloalkyl radical. Illustrative examples of cycloalkyl groups include,but are not limited to the following moieties: C₃₋₆ carbocyclyl groupsinclude, without limitation, cyclopropyl (C₃), cyclobutyl (C₄),cyclopentyl (C₅), cyclopentenyl (C₅), cyclohexyl (C₆), cyclohexenyl(C₆), cyclohexadienyl (C₆), and the like. Examples of C₃₋₈ carbocyclylgroups include the aforementioned C₃₋₆ carbocyclyl groups as well ascycloheptyl (C₇), cycloheptadienyl (C₇), cycloheptatrienyl (C₇),cyclooctyl (C₈), bicyclo[2.2.1]heptanyl, bicyclo[2.2.2]octanyl, and thelike. Examples of C₃₋₁₀ carbocyclyl groups include the aforementionedC₃₋₈ carbocyclyl groups as well as octahydro-1H-indenyl,decahydronaphthalenyl, spiro[4.5]decanyl, and the like. Unless statedotherwise in the specification, a cycloalkyl group is optionallysubstituted by one or more substituents which independently include:acyl, alkyl, alkenyl, alkynyl, alkoxy, alkylaryl, cycloalkyl, aralkyl,aryl, aryloxy, amino, amido, amidino, imino, azide, carbonate,carbamate, carbonyl, heteroalkyl, heteroaryl, heteroarylalkyl,heterocycloalkyl, hydroxy, cyano, halo, haloalkoxy, haloalkyl, ester,ether, mercapto, thio, alkylthio, arylthio, thiocarbonyl, nitro, oxo,phosphate, phosphonate, phosphinate, silyl, sulfinyl, sulfonyl,sulfonamidyl, sulfoxyl, sulfonate, urea, —Si(R^(a))₃, —OR^(a), —SR^(a),—OC(O)—R^(a), —N(R^(a))₂, —C(O)R^(a), —C(O)OR^(a), —OC(O)N(R^(a))₂,—C(O)N(R^(a))₂, —N(R^(a))C(O)OR^(a), —N(R^(a))C(O)R^(a),—N(R^(a))C(O)N(R^(a))₂, —N(R^(a))C(NR^(a))N(R^(a))₂,—N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2), —S(O)_(t)OR^(a) (where t is1 or 2), —S(O)_(t)N(R^(a)) ₂ (where t is 1 or 2), or —O—P(═O)(OR^(a))₂,where each R^(a) is independently hydrogen, alkyl, haloalkyl,carbocyclyl, carbocyclylalkyl, aryl, aralkyl, heterocycloalkyl,heterocycloalkylalkyl, heteroaryl, or heteroarylalkyl, and each of thesemoieties can be optionally substituted as defined herein. In oneembodiment, unless stated otherwise, “cycloalkyl” or “carbocyclyl” alsoincludes ring systems wherein the cycloalkyl or carbocyclyl ring, asdefined above, is fused with one or more aryl or heteroaryl groupswherein the point of attachment to the parent molecular structure is onthe cycloalkyl or carbocyclyl ring.

“Cycloalkyl-alkyl” refers to a -(cycloalkyl)alkyl radical wherecycloalkyl and alkyl are as disclosed herein and which are optionallysubstituted by one or more of the substituents described as suitablesubstituents for cycloalkyl and alkyl respectively. The“cycloalkyl-alkyl” is bonded to the parent molecular structure throughthe cycloalkyl group. The terms “cycloalkyl-alkenyl” and“cycloalkyl-alkynyl” mirror the above description of “cycloalkyl-alkyl”wherein the term “alkyl” is replaced with “alkenyl” or “alkynyl”respectively, and “alkenyl” or “alkynyl” are as described herein.

“Cycloalkyl-heterocycloalkyl” refers to a -(cycloalkyl)heterocyclylalkylradical where cycloalkyl and heterocycloalkyl are as disclosed hereinand which are optionally substituted by one or more of the substituentsdescribed as suitable substituents for heterocycloalkyl and cycloalkylrespectively. The “cycloalkyl-heterocycloalkyl” is bonded to the parentmolecular structure through the cycloalkyl group.

“Cycloalkyl-heteroaryl” refers to a -(cycloalkyl)heteroaryl radicalwhere cycloalkyl and heteroaryl are as disclosed herein and which areoptionally substituted by one or more of the substituents described assuitable substituents for heteroaryl and cycloalkyl respectively. The“cycloalkyl-heteroaryl” is bonded to the parent molecular structurethrough the cycloalkyl group.

As used herein, a “covalent bond” or “direct bond” refers to a singlebond joining two groups.

“Ester” refers to a radical of formula —COOR, where R is selected fromalkyl, alkenyl, alkynyl, haloalkyl, heteroalkyl (bonded through a chaincarbon), cycloalkyl, cycloalkylalkyl, aryl, aralkyl, heterocycloalkyl(bonded through a ring carbon), heterocycloalkylalkyl, heteroaryl(bonded through a ring carbon), and heteroarylalkyl. Any amine, hydroxy,or carboxyl side chain on the compounds described herein can beesterified. The procedures and specific groups to make such esters areknown to those of skill in the art and can readily be found in referencesources such as Greene and Wuts, Protective Groups in Organic Synthesis,4th Ed., John Wiley & Sons, New York, N.Y., 2006, which is incorporatedherein by reference in its entirety. Unless stated otherwise in thespecification, an ester group can be optionally substituted by one ormore substituents which independently include: acyl, alkyl, alkenyl,alkynyl, alkoxy, alkylaryl, cycloalkyl, aralkyl, aryl, aryloxy, amino,amido, amidino, imino, azide, carbonate, carbamate, carbonyl,heteroalkyl, heteroaryl, heteroarylalkyl, heterocycloalkyl, hydroxy,cyano, halo, haloalkoxy, haloalkyl, ester, ether, mercapto, thio,alkylthio, arylthio, thiocarbonyl, nitro, oxo, phosphate, phosphonate,phosphinate, silyl, sulfinyl, sulfonyl, sulfonamidyl, sulfoxyl,sulfonate, urea, —Si(R^(a))₃, —OR^(a), —OC(O)—R^(a), —N(R^(a))₂,—C(O)R^(a), —C(O)OR^(a), —OC(O)N(R^(a))₂, —C(O)N(R^(a))₂,—N(R^(a))C(O)OR^(a), —N(R^(a))C(O)R^(a), —N(R^(a))C(O)N(R^(a))₂,—N(R^(a))C(NR^(a))N(R^(a))₂, —N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2),—S(O)_(t)OR^(a) (where t is 1 or 2), —S(O)_(t)N(R^(a))₂ (where t is 1 or2), or —O—P(═O)(OR^(a))₂, where each R^(a) is independently hydrogen,alkyl, haloalkyl, carbocyclyl, carbocyclylalkyl, aryl, aralkyl,heterocycloalkyl, heterocycloalkylalkyl, heteroaryl, or heteroarylalkyl,and each of these moieties can be optionally substituted as definedherein.

“Ether” refers to a —R^(b)—O—R^(b) radical where each R^(b) isindependently selected from alkyl, alkenyl, alkynyl, haloalkyl,heteroalkyl (bonded through a chain carbon), cycloalkyl,cycloalkylalkyl, aryl, aralkyl, heterocycloalkyl (bonded through a ringcarbon), heterocycloalkylalkyl, heteroaryl (bonded through a ringcarbon), and heteroarylalkyl, unless stated otherwise in thespecification, each of which moiety can itself be optionally substitutedas described herein.

“Halo”, “halide”, or, alternatively, “halogen” means fluoro, chloro,bromo, or iodo. The terms “haloalkyl,” “haloalkenyl,” “haloalkynyl” and“haloalkoxy” include alkyl, alkenyl, alkynyl and alkoxy structures thatare substituted with one or more halo groups or with combinationsthereof. For example, the terms “fluoroalkyl” and “fluoroalkoxy” includehaloalkyl and haloalkoxy groups, respectively, in which the halo isfluorine, such as, but not limited to, trifluoromethyl, difluoromethyl,2,2,2-trifluoroethyl, 1-fluoromethyl-2-fluoroethyl, and the like. Eachof the alkyl, alkenyl, alkynyl and alkoxy groups are as defined hereinand can be optionally further substituted as defined herein.

“Heteroalkyl”, “heteroalkenyl” and “heteroalkynyl” include alkyl,alkenyl and alkynyl radicals, respectively, which have one or moreskeletal chain atoms selected from an atom other than carbon, e.g.,oxygen, nitrogen, sulfur, and phosphorus, or combinations thereof. Anumerical range can be given, e.g., C₁-C₄ heteroalkyl which refers tothe chain length in total, which in this example can be up to 4 atomslong. For example, a —CH₂OCH₂CH₃ radical is referred to as a “C4”heteroalkyl, which includes the heteroatom center in the atom chainlength description. Connection to the parent molecular structure can bethrough either a heteroatom or a carbon in the heteroalkyl chain. Forexample, an N-containing heteroalkyl moiety refers to a group in whichat least one of the skeletal atoms is a nitrogen atom. One or moreheteroatom(s) in the heteroalkyl radical can be optionally oxidized. Oneor more nitrogen atoms, if present, can also be optionally quaternized.For example, heteroalkyl also includes skeletal chains substituted withone or more nitrogen oxide (—O—) substituents. Exemplary heteroalkylgroups include, without limitation, ethers such as methoxyethanyl(—CH₂CH₂OCH₃), ethoxymethanyl (—CH₂OCH₂CH₃), (methoxymethoxy)ethanyl(—CH₂CH₂—OCH₂OCH₃), (methoxymethoxy)methanyl (—CH₂OCH₂OCH₃), and(methoxyethoxy)methanyl (—CH₂OCH₂CH₂OCH₃), and the like; amines such as—CH₂CH₂NHCH₃, CH₂CH₂N(CH₃)₂, —CH₂NHCH₂CH₃, —CH₂N(CH₂CH₃)(CH₃), and thelike. Heteroalkyl, heteroalkenyl, and heteroalkynyl groups can each beoptionally substituted by one or more substituents which independentlyinclude: acyl, alkyl, alkenyl, alkynyl, alkoxy, alkylaryl, cycloalkyl,aralkyl, aryl, aryloxy, amino, amido, amidino, imino, azide, carbonate,carbamate, carbonyl, heteroalkyl, heteroaryl, heteroarylalkyl,heterocycloalkyl, hydroxy, cyano, halo, haloalkoxy, haloalkyl, ester,ether, mercapto, thio, alkylthio, arylthio, thiocarbonyl, nitro, oxo,phosphate, phosphonate, phosphinate, silyl, sulfinyl, sulfonyl,sulfonamidyl, sulfoxyl, sulfonate, urea, —Si(R^(a))₃, —OR^(a), —SR^(a),—OC(O)—R^(a), —N(R^(a))₂, —C(O)R^(a), —C(O)OR^(a), —OC(O)N(R^(a))₂,—C(O)N(R^(a))₂, —N(R^(a))C(O)OR^(a), —N(R^(a))C(O)R^(a),—N(R^(a))C(O)N(R^(a))₂, —N(R^(a))C(NR^(a))N(R^(a))₂,—N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2), —S(O)_(t)OR^(a) (where t is1 or 2), —S(O)_(t)N(R^(a))₂ (where t is 1 or 2), or —O—P(═O)(OR^(a))₂,where each R^(a) is independently hydrogen, alkyl, haloalkyl,carbocyclyl, carbocyclylalkyl, aryl, aralkyl, heterocycloalkyl,heterocycloalkylalkyl, heteroaryl, or heteroarylalkyl, and each of thesemoieties can be optionally substituted as defined herein.

“Heteroalkyl-aryl” refers to a -(heteroalkyl)aryl radical whereheteroalkyl and aryl are as disclosed herein and which are optionallysubstituted by one or more of the substituents described as suitablesubstituents for heteroalkyl and aryl respectively. The“heteroalkyl-aryl” is bonded to the parent molecular structure throughan atom of the heteroalkyl group.

“Heteroalkyl-heteroaryl” refers to a -(heteroalkyl)heteroaryl radicalwhere heteroalkyl and heteroaryl are as disclosed herein and which areoptionally substituted by one or more of the substituents described assuitable substituents for heteroalkyl and heteroaryl respectively. The“heteroalkyl-heteroaryl” is bonded to the parent molecular structurethrough an atom of the heteroalkyl group.

“Heteroalkyl-heterocycloalkyl” refers to a-(heteroalkyl)heterocycloalkyl radical where heteroalkyl andheterocycloalkyl are as disclosed herein and which are optionallysubstituted by one or more of the substituents described as suitablesubstituents for heteroalkyl and heterocycloalkyl respectively. The“heteroalkyl-heterocycloalkyl” is bonded to the parent molecularstructure through an atom of the heteroalkyl group.

“Heteroalkyl-cycloalkyl” refers to a -(heteroalkyl)cycloalkyl radicalwhere heteroalkyl and cycloalkyl are as disclosed herein and which areoptionally substituted by one or more of the substituents described assuitable substituents for heteroalkyl and cycloalkyl respectively. The“heteroalkyl-cycloalkyl” is bonded to the parent molecular structurethrough an atom of the heteroalkyl group.

“Heteroaryl”, or alternatively, “heteroaromatic”, refers to a radical ofa 5- to 18-membered monocyclic or polycyclic (e.g., bicyclic ortricyclic) aromatic ring system (e.g., having 6, 10 or 14 π electronsshared in a cyclic array) having ring carbon atoms and 1 to 6 ringheteroatoms provided in the aromatic ring system, wherein eachheteroatom is independently selected from nitrogen, oxygen, phosphorous,and sulfur (“5- to 18-membered heteroaryl”). Heteroaryl polycyclic ringsystems can include one or more heteroatoms in one or more rings.Whenever it appears herein, a numerical range such as “5 to 18” refersto each integer in the given range; e.g., “5 to 18 ring atoms” meansthat the heteroaryl group can consist of 5 ring atoms, 6 ring atoms, 7ring atoms, 8 ring atoms, 9 ring atoms, 10 ring atoms, etc., up to andincluding 18 ring atoms. In one embodiment, bivalent radicals derivedfrom univalent heteroaryl radicals whose names end in “-yl” by removalof one hydrogen atom from the atom with the free valence are named byadding “-idene” to the name of the corresponding univalent radical,e.g., a pyridyl group with two points of attachment is a pyridylidene.

For example, an N-containing “heteroaromatic” or “heteroaryl” moietyrefers to an aromatic group in which at least one of the skeletal atomsof the ring is a nitrogen atom. One or more heteroatom(s) in theheteroaryl radical can be optionally oxidized. One or more nitrogenatoms, if present, can also be optionally quaternized. Heteroaryl alsoincludes ring systems substituted with one or more nitrogen oxide (—O—)substituents, such as pyridinyl N-oxides. The heteroaryl is attached tothe parent molecular structure through any atom of the ring(s).

“Heteroaryl” also includes ring systems wherein the heteroaryl ring, asdefined above, is fused with one or more aryl groups wherein the pointof attachment to the parent molecular structure is either on the aryl oron the heteroaryl ring, or wherein the heteroaryl ring, as definedabove, is fused with one or more cycloalkyl or heterocyclyl groupswherein the point of attachment to the parent molecular structure is onthe heteroaryl ring. For polycyclic heteroaryl groups wherein one ringdoes not contain a heteroatom (e.g., indolyl, quinolinyl, carbazolyl andthe like), the point of attachment to the parent molecular structure canbe on either the ring bearing a heteroatom (e.g., 2-indolyl) or the ringthat does not contain a heteroatom (e.g., 5-indolyl). In someembodiments, a heteroaryl group is a 5 to 10 membered aromatic ringsystem having ring carbon atoms and 1 to 4 ring heteroatoms provided inthe aromatic ring system, wherein each heteroatom is independentlyselected from nitrogen, oxygen, phosphorous, and sulfur (“5- to10-membered heteroaryl”). In some embodiments, a heteroaryl group is a5- to 8-membered aromatic ring system having ring carbon atoms and 1 to4 ring heteroatoms provided in the aromatic ring system, wherein eachheteroatom is independently selected from nitrogen, oxygen, phosphorous,and sulfur (“5- to 8-membered heteroaryl”). In some embodiments, aheteroaryl group is a 5- to 6-membered aromatic ring system having ringcarbon atoms and 1 to 4 ring heteroatoms provided in the aromatic ringsystem, wherein each heteroatom is independently selected from nitrogen,oxygen, phosphorous, and sulfur (“5- to 6-membered heteroaryl”). In someembodiments, the 5- to 6-membered heteroaryl has 1 to 3 ring heteroatomsindependently selected from nitrogen, oxygen, phosphorous, and sulfur.In some embodiments, the 5- to 6-membered heteroaryl has 1 to 2 ringheteroatoms independently selected from nitrogen, oxygen, phosphorous,and sulfur. In some embodiments, the 5- to 6-membered heteroaryl has 1ring heteroatom selected from nitrogen, oxygen, phosphorous, and sulfur.

Examples of heteroaryls include, but are not limited to, azepinyl,acridinyl, benzimidazolyl, benzindolyl, 1,3-benzodioxolyl, benzofuranyl,benzooxazolyl, benzo[d]thiazolyl, benzothiadiazolyl,benzo[b][1,4]dioxepinyl, benzo[b][1,4]oxazinyl, 1,4-benzodioxanyl,benzonaphthofuranyl, benzoxazolyl, benzodioxolyl, benzodioxinyl,benzoxazolyl, benzopyranyl, benzopyranonyl, benzofuranyl,benzofuranonyl, benzofurazanyl, benzothiazolyl, benzothienyl(benzothiophenyl), benzothieno[3,2-d]pyrimidinyl, benzotriazolyl,benzo[4,6]imidazo[1,2-a]pyridinyl, carbazolyl, cinnolinyl,cyclopenta[d]pyrimidinyl,6,7-dihydro-5H-cyclopenta[4,5]thieno[2,3-d]pyrimidinyl,5,6-dihydrobenzo[h]quinazolinyl, 5,6-dihydrobenzo[h]cinnolinyl,6,7-dihydro-5H-benzo[6,7]cyclohepta[1,2-c]pyridazinyl, dibenzofuranyl,dibenzothiophenyl, furanyl, furazanyl, furanonyl, furo[3,2-c]pyridinyl,5,6,7,8,9,10-hexahydrocycloocta[d]pyrimidinyl,5,6,7,8,9,10-hexahydrocycloocta[d]pyridazinyl,5,6,7,8,9,10-hexahydrocycloocta[d]pyridinyl, isothiazolyl, imidazolyl,indazolyl, indolyl, indazolyl, isoindolyl, indolinyl, isoindolinyl,isoquinolyl, indolizinyl, isoxazolyl,5,8-methano-5,6,7,8-tetrahydroquinazolinyl, naphthyridinyl,1,6-naphthyridinonyl, oxadiazolyl, 2-oxoazepinyl, oxazolyl, oxiranyl,5,6,6a,7,8,9,10,10a-octahydrobenzo[h]quinazolinyl, 1-phenyl-1H-pyrrolyl,phenazinyl, phenothiazinyl, phenoxazinyl, phthalazinyl, pteridinyl,purinyl, pyranyl, pyrrolyl, pyrazolyl, pyrazolo[3,4-d]pyrimidinyl,pyridinyl, pyrido[3,2-d]pyrimidinyl, pyrido[3,4-d]pyrimidinyl,pyrazinyl, pyrimidinyl, pyridazinyl, pyrrolyl, quinazolinyl,quinoxalinyl, quinolinyl, isoquinolinyl, tetrahydroquinolinyl,5,6,7,8-tetrahydroquinazolinyl,5,6,7,8-tetrahydrobenzo[4,5]thieno[2,3-d]pyrimidinyl,6,7,8,9-tetrahydro-5H-cyclohepta[4,5]thieno[2,3-d] pyrimidinyl,5,6,7,8-tetrahydropyrido[4,5-c]pyridazinyl, thiazolyl, thiadiazolyl,thiapyranyl, triazolyl, tetrazolyl, triazinyl, thieno[2,3-d]pyrimidinyl,thieno[3,2-d]pyrimidinyl, thieno[2,3-c]pridinyl, and thiophenyl (i.e.,thienyl).

Unless stated otherwise in the specification, a heteroaryl moiety isoptionally substituted by one or more substituents which independentlyinclude: acyl, alkyl, alkenyl, alkynyl, alkoxy, alkylaryl, cycloalkyl,aralkyl, aryl, aryloxy, amino, amido, amidino, imino, azide, carbonate,carbamate, carbonyl, heteroalkyl, heteroaryl, heteroarylalkyl,heterocycloalkyl, hydroxy, cyano, halo, haloalkoxy, haloalkyl, ester,ether, mercapto, thio, alkylthio, arylthio, thiocarbonyl, nitro, oxo,phosphate, phosphonate, phosphinate, silyl, sulfinyl, sulfonyl,sulfonamidyl, sulfoxyl, sulfonate, urea, —Si(R^(a))₃, —OR^(a), —SR^(a),—OC(O)—R^(a), —N(R^(a))₂, —C(O)R^(a), —C(O)OR^(a), —OC(O)N(R^(a))₂,—C(O)N(R^(a))₂, —N(R^(a))C(O)OR^(a), —N(R^(a))C(O)R^(a),—N(R^(a))C(O)N(R^(a))₂, —N(R^(a))C(NR^(a))N(R^(a))₂,—N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2), —S(O)_(t)OR^(a) (where t is1 or 2), —S(O)_(t)N(R^(a))₂ (where t is 1 or 2), or —O—P(═O)(OR^(a))₂,where each R^(a) is independently hydrogen, alkyl, haloalkyl,carbocyclyl, carbocyclylalkyl, aryl, aralkyl, heterocycloalkyl,heterocycloalkylalkyl, heteroaryl, or heteroarylalkyl, and each of thesemoieties can be optionally substituted as defined herein.

“Heteroaryl-alkyl” refers to a -(heteroaryl)alkyl radical whereheteroaryl and alkyl are as disclosed herein and which are optionallysubstituted by one or more of the substituents described as suitablesubstituents for heteroaryl and alkyl respectively. The“heteroaryl-alkyl” is bonded to the parent molecular structure throughany atom of the heteroaryl group.

“Heteroaryl-heterocycloalkyl” refers to an -(heteroaryl)heterocycloalkylradical where heteroaryl and heterocycloalkyl are as disclosed hereinand which are optionally substituted by one or more of the substituentsdescribed as suitable substituents for heteroaryl and heterocycloalkylrespectively. The “heteroaryl-heterocycloalkyl” is bonded to the parentmolecular structure through an atom of the heteroaryl group.

“Heteroaryl-cycloalkyl” refers to an -(heteroaryl)cycloalkyl radicalwhere heteroaryl and cycloalkyl are as disclosed herein and which areoptionally substituted by one or more of the substituents described assuitable substituents for heteroaryl and cycloalkyl respectively. The“heteroaryl-cycloalkyl” is bonded to the parent molecular structurethrough a carbon atom of the heteroaryl group.

“Heterocyclyl”, “heterocycloalkyl” or ‘heterocarbocyclyl” each refer toany 3- to 18-membered non-aromatic radical monocyclic or polycyclicmoiety comprising at least one ring heteroatom selected from nitrogen,oxygen, phosphorous, and sulfur. A heterocyclyl group can be amonocyclic, bicyclic, tricyclic or tetracyclic ring system, wherein thepolycyclic ring systems can be a fused, bridged or spiro ring system.Heterocyclyl polycyclic ring systems can include one or more heteroatomsin one or more rings. A heterocyclyl group can be saturated or partiallyunsaturated. Partially unsaturated heterocycloalkyl groups can be termed“heterocycloalkenyl” if the heterocyclyl contains at least one doublebond, or “heterocycloalkynyl” if the heterocyclyl contains at least onetriple bond. Whenever it appears herein, a numerical range such as “5 to18” refers to each integer in the given range; e.g., “5 to 18 ringatoms” means that the heterocyclyl group can consist of 5 ring atoms, 6ring atoms, 7 ring atoms, 8 ring atoms, 9 ring atoms, 10 ring atoms,etc., up to and including 18 ring atoms. In one embodiment, bivalentradicals derived from univalent heterocyclyl radicals whose names end in“-yl” by removal of one hydrogen atom from the atom with the freevalence are named by adding “-idene” to the name of the correspondingunivalent radical, e.g., a piperidyl group with two points of attachmentis a piperidylidene.

An N-containing heterocyclyl moiety refers to an non-aromatic group inwhich at least one of the ring atoms is a nitrogen atom. Theheteroatom(s) in the heterocyclyl radical can be optionally oxidized.One or more nitrogen atoms, if present, can be optionally quaternized.Heterocyclyl also includes ring systems substituted with one or morenitrogen oxide (—O—) substituents, such as piperidinyl N-oxides. Theheterocyclyl is attached to the parent molecular structure through anyatom of any of the ring(s).

“Heterocyclyl” also includes ring systems wherein the heterocyclyl ring,as defined above, is fused with one or more carbocyclyl groups whereinthe point of attachment is either on the carbocyclyl or heterocyclylring, or ring systems wherein the heterocyclyl ring, as defined above,is fused with one or more aryl or heteroaryl groups, wherein the pointof attachment to the parent molecular structure is on the heterocyclylring. In some embodiments, a heterocyclyl group is a 3- to 10-memberednon-aromatic ring system having ring carbon atoms and 1 to 4 ringheteroatoms, wherein each heteroatom is independently selected fromnitrogen, oxygen, phosphorous, and sulfur (“3- to 10-memberedheterocyclyl”). In some embodiments, a heterocyclyl group is a 5- to8-membered non-aromatic ring system having ring carbon atoms and 1 to 4ring heteroatoms, wherein each heteroatom is independently selected fromnitrogen, oxygen, phosphorous, and sulfur (“5- to 8-memberedheterocyclyl”). In some embodiments, a heterocyclyl group is a 5- to6-membered non-aromatic ring system having ring carbon atoms and 1 to 4ring heteroatoms, wherein each heteroatom is independently selected fromnitrogen, oxygen, phosphorous, and sulfur (“5- to 6-memberedheterocyclyl”). In some embodiments, the 5- to 6-membered heterocyclylhas 1 to 3 ring heteroatoms independently selected from nitrogen,oxygen, phosphorous, and sulfur. In some embodiments, the 5- to6-membered heterocyclyl has 1 to 2 ring heteroatoms independentlyselected from nitrogen, oxygen, phosphorous, and sulfur. In someembodiments, the 5- to 6-membered heterocyclyl has 1 ring heteroatomselected from nitrogen, oxygen, phosphorous, and sulfur.

Exemplary 3-membered heterocyclyls containing 1 heteroatom include,without limitation, azirdinyl, oxiranyl, thiorenyl. Exemplary 4-memberedheterocyclyls containing 1 heteroatom include, without limitation,azetidinyl, oxetanyl and thietanyl. Exemplary 5-membered heterocyclylscontaining 1 heteroatom include, without limitation, tetrahydrofuranyl,dihydrofuranyl, tetrahydrothiophenyl, dihydrothiophenyl, pyrrolidinyl,dihydropyrrolyl and pyrrolyl-2,5-dione. Exemplary 5-memberedheterocyclyls containing 2 heteroatoms include, without limitation,dioxolanyl, oxathiolanyl and dithiolanyl. Exemplary 5-memberedheterocyclyls containing 3 heteroatoms include, without limitation,triazolinyl, oxadiazolinyl, and thiadiazolinyl. Exemplary 6-memberedheterocyclyl groups containing 1 heteroatom include, without limitation,piperidinyl, tetrahydropyranyl, dihydropyridinyl, and thianyl. Exemplary6-membered heterocyclyl groups containing 2 heteroatoms include, withoutlimitation, piperazinyl, morpholinyl, dithianyl, dioxanyl, andtriazinanyl. Exemplary 7-membered heterocyclyl groups containing 1heteroatom include, without limitation, azepanyl, oxepanyl andthiepanyl. Exemplary 8-membered heterocyclyl groups containing 1heteroatom include, without limitation, azocanyl, oxecanyl andthiocanyl. Exemplary bicyclic heterocyclyl groups include, withoutlimitation, indolinyl, isoindolinyl, dihydrobenzofuranyl,dihydrobenzothienyl, tetrahydrobenzothienyl, tetrahydrobenzofuranyl,tetrahydroindolyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl,decahydroquinolinyl, decahydroisoquinolinyl, octahydrochromenyl,octahydroisochromenyl, decahydronaphthyridinyl,decahydro-1,8-naphthyridinyl, octahydropyrrolo[3,2-b]pyrrole, indolinyl,phthalimidyl, naphthalimidyl, chromanyl, chromenyl,1H-benzo[e][1,4]diazepinyl, 1,4,5,7-tetrahydro-pyrano[3,4-b]pyrrolyl,5,6-dihydro-4H-furo[3,2-b]pyrrolyl, 6,7-dihydro-5H-furo[3,2-b]pyranyl,5,7-dihydro-4H-thieno[2,3-c]pyranyl,2,3-dihydro-1H-pyrrolo[2,3-b]pyridinyl, 2,3-dihydrofuro[2,3-b]pyridinyl,4,5,6,7-tetrahydro-1H-pyrrolo[2,3-b]pyridinyl,4,5,6,7-tetrahydrofuro[3,2-c]pyridinyl,4,5,6,7-tetrahydrothieno[3,2-b]pyridinyl,1,2,3,4-tetrahydro-1,6-naphthyridinyl, and the like.

Unless stated otherwise, heterocyclyl moieties are optionallysubstituted by one or more substituents which independently include:acyl, alkyl, alkenyl, alkynyl, alkoxy, alkylaryl, cycloalkyl, aralkyl,aryl, aryloxy, amino, amido, amidino, imino, azide, carbonate,carbamate, carbonyl, heteroalkyl, heteroaryl, heteroarylalkyl,heterocycloalkyl, hydroxy, cyano, halo, haloalkoxy, haloalkyl, ester,ether, mercapto, thio, alkylthio, arylthio, thiocarbonyl, nitro, oxo,phosphate, phosphonate, phosphinate, silyl, sulfinyl, sulfonyl,sulfonamidyl, sulfoxyl, sulfonate, urea, —Si(R^(a))₃, —OR^(a), —SR^(a),—OC(O)—R^(a), —N(R^(a))₂, —C(O)R^(a), —C(O)OR^(a), —OC(O)N(R^(a))₂,—C(O)N(R^(a))₂, —N(R^(a))C(O)OR^(a), —N(R^(a))C(O)R^(a),—N(R^(a))C(O)N(R^(a))₂, —N(R^(a))C(NR^(a))(R^(a))₂,—N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2), —S(O)_(t)OR^(a) (where t is1 or 2), —S(O)_(t)N(R^(a))₂ (where t is 1 or 2), or —O—P(═O)(OR^(a))₂,where each R^(a) is independently hydrogen, alkyl, haloalkyl,carbocyclyl, carbocyclylalkyl, aryl, aralkyl, heterocycloalkyl,heterocycloalkylalkyl, heteroaryl, or heteroarylalkyl, and each of thesemoieties can be optionally substituted as defined herein.

“Heterocyclyl-alkyl” refers to a -(heterocyclyl)alkyl radical whereheterocyclyl and alkyl are as disclosed herein and which are optionallysubstituted by one or more of the substituents described as suitablesubstituents for heterocyclyl and alkyl respectively. The“heterocyclyl-alkyl” is bonded to the parent molecular structure throughany atom of the heterocyclyl group. The terms “heterocyclyl-alkenyl” and“heterocyclyl-alkynyl” mirror the above description of“heterocyclyl-alkyl” wherein the term “alkyl” is replaced with “alkenyl”or “alkynyl” respectively, and “alkenyl” or “alkynyl” are as describedherein.

“Imino” refers to the “—C(═N—R^(b))—R^(b)” radical where each R^(b) isindependently selected from hydrogen, alkyl, alkenyl, alkynyl,haloalkyl, heteroalkyl (bonded through a chain carbon), cycloalkyl,cycloalkylalkyl, aryl, aralkyl, heterocycloalkyl (bonded through a ringcarbon), heterocycloalkylalkyl, heteroaryl (bonded through a ringcarbon), and heteroarylalkyl, unless stated otherwise in thespecification, each of which moiety can itself be optionally substitutedas described herein.

“Moiety” refers to a specific segment or functional group of a molecule.Chemical moieties are often recognized chemical entities embedded in orappended to a molecule.

“Nitro” refers to the —NO₂ radical.

“Oxa” refers to the —O— radical.

“Oxo” refers to the ═O radical.

“Phosphate” refers to a —O—P(═O)(OR^(b))₂ radical, where each R^(b) isindependently selected from hydrogen, alkyl, alkenyl, alkynyl,haloalkyl, heteroalkyl (bonded through a chain carbon), cycloalkyl,cycloalkylalkyl, aryl, aralkyl, heterocycloalkyl (bonded through a ringcarbon), heterocycloalkylalkyl, heteroaryl (bonded through a ringcarbon), and heteroarylalkyl, unless stated otherwise in thespecification, each of which moiety can itself be optionally substitutedas described herein. In some embodiments, when R^(a) is hydrogen anddepending on the pH, the hydrogen can be replaced by an appropriatelycharged counter ion.

“Phosphonate” refers to a —O—P(═O)(R^(b))(OR^(b)) radical, where eachR^(b) is independently selected from hydrogen, alkyl, alkenyl, alkynyl,haloalkyl, heteroalkyl (bonded through a chain carbon), cycloalkyl,cycloalkylalkyl, aryl, aralkyl, heterocycloalkyl (bonded through a ringcarbon), heterocycloalkylalkyl, heteroaryl (bonded through a ringcarbon) and heteroarylalkyl, unless stated otherwise in thespecification, each of which moiety can itself be optionally substitutedas described herein. In some embodiments, when R^(a) is hydrogen anddepending on the pH, the hydrogen can be replaced by an appropriatelycharged counter ion.

“Phosphinate” refers to a —P(═O)(R^(b))(OR^(b)) radical, where eachR^(b) is independently selected from hydrogen, alkyl, alkenyl, alkynyl,haloalkyl, heteroalkyl (bonded through a chain carbon), cycloalkyl,cycloalkylalkyl, aryl, aralkyl, heterocycloalkyl (bonded through a ringcarbon), heterocycloalkylalkyl, heteroaryl (bonded through a ringcarbon), and heteroarylalkyl, unless stated otherwise in thespecification, each of which moiety can itself be optionally substitutedas described herein. In some embodiments, when R^(a) is hydrogen anddepending on the pH, the hydrogen can be replaced by an appropriatelycharged counter ion.

A “leaving group or atom” is any group or atom that will, under thereaction conditions, cleave from the starting material, thus promotingreaction at a specified site. Suitable non-limiting examples of suchgroups, unless otherwise specified, include halogen atoms, mesyloxy,p-nitrobenzensulphonyloxy, trifluoromethyloxy, and tosyloxy groups.

“Protecting group” has the meaning conventionally associated with it inorganic synthesis, e.g., a group that selectively blocks one or morereactive sites in a multifunctional compound such that a chemicalreaction can be carried out selectively on another unprotected reactivesite and such that the group can readily be removed after the selectivereaction is complete. A variety of protecting groups are disclosed, forexample, in T. H. Greene and P. G. M. Wuts, Protective Groups in OrganicSynthesis, Fourth Edition, John Wiley & Sons, New York (2006),incorporated herein by reference in its entirety. For example, a hydroxyprotected form is where at least one of the hydroxy groups present in acompound is protected with a hydroxy protecting group. Likewise, aminesand other reactive groups can similarly be protected.

As used herein, the terms “substituted” or “substitution” mean that atleast one hydrogen present on a group atom (e.g., a carbon or nitrogenatom) is replaced with a permissible substituent, e.g., a substituentwhich upon substitution for the hydrogen results in a stable compound,e.g., a compound which does not spontaneously undergo transformationsuch as by rearrangement, cyclization, elimination, or other reaction.Unless otherwise indicated, a “substituted” group can have a substituentat one or more substitutable positions of the group, and when more thanone position in any given structure is substituted, the substituent iseither the same or different at each position. Substituents can includeone or more group(s) individually and independently selected from acyl,alkyl, alkenyl, alkynyl, alkoxy, alkylaryl, cycloalkyl, aralkyl, aryl,aryloxy, amino, amido, azide, carbonate, carbonyl, heteroalkyl,heteroaryl, heteroarylalkyl, heterocycloalkyl, hydroxy, cyano, halo,haloalkoxy, haloalkyl, ester, mercapto, thio, alkylthio, arylthio,thiocarbonyl, nitro, oxo, phosphate, phosphonate, phosphinate, silyl,sulfinyl, sulfonyl, sulfonamidyl, sulfoxyl, sulfonate, urea,—Si(R^(a))₃, —OR^(a), —SR^(a), —OC(O)—R^(a), —N(R^(a))₂, —C(O)R^(a),—C(O)OR^(a), —OC(O)N(R^(a))₂, —C(O)N(R^(a))₂, —N(R^(a))C(O)OR^(a),—N(R^(a))C(O)R^(a), —N(R^(a))C(O)N(R^(a))₂, —N(R^(a))C(NR^(a))N(R^(a))₂,—N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2), —S(O)_(t)OR^(a) (where t is1 or 2), —S(O)_(t)N(R^(a))₂ (where t is 1 or 2), and —O—P(═O)(OR^(a))₂,where each R^(a) is independently hydrogen, alkyl, haloalkyl,carbocyclyl, carbocyclylalkyl, aryl, aralkyl, heterocycloalkyl,heterocycloalkylalkyl, heteroaryl, or heteroarylalkyl, and each of thesemoieties can be optionally substituted as defined herein. For example, acycloalkyl substituent can have a halide substituted at one or more ringcarbons, and the like. The protecting groups that can form theprotective derivatives of the above substituents are known to those ofskill in the art and can be found in references such as Greene and Wuts,above.

“Silyl” refers to a —Si(R^(b))₃ radical where each R^(b) isindependently selected from alkyl, alkenyl, alkynyl, haloalkyl,heteroalkyl (bonded through a chain carbon), cycloalkyl,cycloalkylalkyl, aryl, aralkyl, heterocycloalkyl (bonded through a ringcarbon), heterocycloalkylalkyl, heteroaryl (bonded through a ringcarbon), and heteroarylalkyl, unless stated otherwise in thespecification, each of which moiety can itself be optionally substitutedas described herein.

“Sulfanyl”, “sulfide”, and “thio” each refer to the radical —S—R^(b),wherein R^(b) is selected from alkyl, alkenyl, alkynyl, haloalkyl,heteroalkyl (bonded through a chain carbon), cycloalkyl,cycloalkylalkyl, aryl, aralkyl, heterocycloalkyl (bonded through a ringcarbon), heterocycloalkylalkyl, heteroaryl (bonded through a ringcarbon), and heteroarylalkyl, unless stated otherwise in thespecification, each of which moiety can itself be optionally substitutedas described herein. For instance, an “alkylthio” refers to the“alkyl-S—” radical, and “arylthio” refers to the “aryl-S—” radical, eachof which are bound to the parent molecular group through the S atom. Theterms “sulfide”, “thiol”, “mercapto”, and “mercaptan” can also eachrefer to the group —R^(b)SH.

“Sulfinyl” or “sulfoxide” refers to the —S(O)—R^(b) radical, wherein for“sulfinyl”, R^(b) is H, and for “sulfoxide”, R^(b) is selected fromalkyl, alkenyl, alkynyl, haloalkyl, heteroalkyl (bonded through a chaincarbon), cycloalkyl, cycloalkylalkyl, aryl, aralkyl, heterocycloalkyl(bonded through a ring carbon), heterocycloalkylalkyl, heteroaryl(bonded through a ring carbon), and heteroarylalkyl, unless statedotherwise in the specification, each of which moiety can itself beoptionally substituted as described herein.

“Sulfonyl” or “sulfone” refers to the —S(O₂)—R^(b) radical, whereinR^(b) is selected from hydrogen, alkyl, alkenyl, alkynyl, haloalkyl,heteroalkyl (bonded through a chain carbon), cycloalkyl,cycloalkylalkyl, aryl, aralkyl, heterocycloalkyl (bonded through a ringcarbon), heterocycloalkylalkyl, heteroaryl (bonded through a ringcarbon), and heteroarylalkyl, unless stated otherwise in thespecification, each of which moiety can itself be optionally substitutedas described herein.

“Sulfonamidyl” or “sulfonamido” refers to the following radicals:—S(═O)₂—N(R^(b))₂, —N(R^(b))—S(═O)₂—R^(b), —S(═O)₂—N(R^(b))—, or—N(R^(b))—S(═O)₂—, where each R^(b) is independently selected fromhydrogen, alkyl, alkenyl, alkynyl, haloalkyl, heteroalkyl (bondedthrough a chain carbon), cycloalkyl, cycloalkylalkyl, aryl, aralkyl,heterocycloalkyl (bonded through a ring carbon), heterocycloalkylalkyl,heteroaryl (bonded through a ring carbon), and heteroarylalkyl, unlessstated otherwise in the specification, each of which moiety can itselfbe optionally substituted as described herein. The R^(b) groups in—S(═O)₂—N(R^(b))₂ or —N(R^(b))—S(═O)₂—R^(b) can be taken together withthe nitrogen to which they are attached to form a 4-, 5-, 6-, 7-, or8-membered heterocyclyl ring. In some embodiments, the term designates aC₁-C₄ sulfonamido, wherein each R^(b) in the sulfonamido contains 1carbon, 2 carbons, 3 carbons, or 4 carbons total.

“Sulfoxyl” refers to a —S(═O)₂OH radical.

“Sulfonate” refers to a —S(═O)₂—OR^(b) radical, wherein R^(b) isselected from alkyl, alkenyl, alkynyl, haloalkyl, heteroalkyl (bondedthrough a chain carbon), cycloalkyl, cycloalkylalkyl, aryl, aralkyl,heterocycloalkyl (bonded through a ring carbon), heterocycloalkylalkyl,heteroaryl (bonded through a ring carbon), and heteroarylalkyl, unlessstated otherwise in the specification, each of which moiety can itselfbe optionally substituted as described herein.

“Thiocarbonyl” refers to a —(C═S)— radical.

“Urea” refers to a —N(R^(b))—(C═O)—N(R^(b))₂ or—N(R^(b))—(C═O)—N(R^(b))— radical, where each R^(b) is independentlyselected from hydrogen, alkyl, alkenyl, alkynyl, haloalkyl, heteroalkyl(bonded through a chain carbon), cycloalkyl, cycloalkylalkyl, aryl,aralkyl, heterocycloalkyl (bonded through a ring carbon),heterocycloalkylalkyl, heteroaryl (bonded through a ring carbon), andheteroarylalkyl, unless stated otherwise in the specification, each ofwhich moiety can itself be optionally substituted as described herein.

Where substituent groups are specified by their conventional chemicalformulae, written from left to right, they equally encompass thechemically identical substituents that would result from writing thestructure from right to left, e.g., —CH₂O— is equivalent to —OCH₂—.

Compounds

In certain embodiments, provided herein are compounds of Formula (I″) orFormula (A″):

wherein:R¹ is hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,aryl, heteroaryl, —COR², —COOR³, or —CONR⁴R⁵;z is 0, 1, 2, or 3;each instance of R^(3a) is independently hydrogen, alkyl, alkenyl,alkynyl, alkoxyl, halogen, cyano, amino, cycloalkyl, heterocycloalkyl,aryl, or heteroaryl;B is hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,aryl, heteroaryl, —COR², —COOR³, —CONR⁴R⁵, or —Si(R⁶)₃;

-   -   wherein R², R³, R⁴, R⁵, and R⁶ are each, independently,        hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,        aryl, or heteroaryl;        R^(1c) is hydrogen, alkyl, alkenyl, or alkynyl;        R^(2c) is hydrogen, alkyl, alkenyl, or alkynyl;        W^(d) is heteroaryl, cycloalkyl, heterocycloalkyl, or aryl; and        X is CR^(1a) or N;    -   wherein R^(1a) is hydrogen, halo, alkyl, alkenyl, alkynyl, or        CN;    -   wherein each alkyl, alkenyl, or alkynyl is optionally        substituted with one or more halo, OH, alkoxy, NH₂, NH(alkyl),        N(alkyl)₂, COH, CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl),        CON(alkyl)₂, S(O)(alkyl), S(O)₂(alkyl), cycloalkyl,        heterocycloalkyl, aryl or heteroaryl;    -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, alkyl, alkenyl,        alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂, COH,        CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl), CON(alkyl)₂,        S(O)(alkyl), or S(O)₂(alkyl);    -   wherein in Formula (I″), when X is CH, B is unsubstituted        phenyl, and W^(d) is

-   -    then R¹ is not hydrogen, Si(CH₃)₃, CH₂Si(CH₃)₃, methyl,        (CH₂)NH₂, (CH₂)₂NH₂, (CH₂)NHSO₂CH₃, or (CH₂)_(n)NHC(O)R^(1x); n        is 1 or 2; R^(1x) is methyl, C₂ alkene, cyclohexyl, cyclopentyl,        tetrahydrofuranyl, furanyl, or pyrrolidinyl, where the alkene,        cyclohexyl, cyclopentyl, tetrahydrofuranyl, furanyl, or        pyrrolidinyl is optionally substituted with one or two groups        independently selected from oxo and cyano;    -   wherein in Formula (A″), when X is CH, B is unsubstituted        phenyl, and W^(d) is

-   -    then R¹ is not phenyl;        or a pharmaceutically acceptable form thereof.

In one embodiment, B is alkyl, alkenyl, alkynyl, cycloalkyl,heterocycloalkyl, aryl, heteroaryl, —COR², —COOR³, or —CONR⁴R⁵. In oneembodiment, B is alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,aryl, heteroaryl, —COR², —COOR³, or —CONR⁴R⁵, wherein the point ofattachment for the heterocycloalkyl and heteroaryl is a carbon atom.

In one embodiment, R^(1c) is alkyl, alkenyl, or alkynyl.

In one embodiment, R^(1c) is hydrogen. In one embodiment, R^(1c) isalkyl. In one embodiment, R^(1c) is methyl or ethyl. In one embodiment,R^(1c) is methyl. In one embodiment, R^(1c) is ethyl.

In one embodiment, R^(2c) is hydrogen.

In one embodiment, in Formula (I″), X is N and R^(2c) is hydrogen.

In one embodiment, z is 0. In another embodiment, z is 1. In anotherembodiment, z is 2. In another embodiment, z is 3.

In one embodiment, each instance of R^(3a) is independently hydrogen,alkyl, or halogen. In one embodiment, each instance of R^(3a) isindependently hydrogen, methyl, fluoro, chloro, or bromo.

In certain embodiments, provided herein are compounds of Formula (I″) orFormula (A″):

wherein:R¹ is hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,aryl, heteroaryl, —COR², —COOR³, or —CONR⁴R⁵;z is 0, 1, 2, or 3;each instance of R^(3a) is independently hydrogen, alkyl, alkenyl,alkynyl, alkoxyl, halogen, cyano, amino, cycloalkyl, heterocycloalkyl,aryl, or heteroaryl;B is alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl,heteroaryl, —COR², —COOR³, or —CONR⁴R⁵, wherein the point of attachmentfor the heterocycloalkyl and heteroaryl is a carbon atom;

-   -   wherein R², R³, R⁴, and R⁵ are each, independently, hydrogen,        alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, or        heteroaryl;        R^(1c) is alkyl, alkenyl, or alkynyl;        R^(2c) is hydrogen;        W^(d) is heteroaryl, cycloalkyl, heterocycloalkyl, or aryl; and        X is CR^(1a) or N;    -   wherein R^(1a) is hydrogen, halo, alkyl, alkenyl, alkynyl, or        CN;    -   wherein each alkyl, alkenyl, or alkynyl is optionally        substituted with one or more halo, OH, alkoxy, NH₂, NH(alkyl),        N(alkyl)₂, COH, CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl),        CON(alkyl)₂, S(O)(alkyl), S(O)₂(alkyl), cycloalkyl,        heterocycloalkyl, aryl or heteroaryl;    -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, alkyl, alkenyl,        alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂, COH,        CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl), CON(alkyl)₂,        S(O)(alkyl), or S(O)₂(alkyl);    -   wherein in Formula (I″), when X is CH, B is unsubstituted        phenyl, and W^(d) is

-   -    then R¹ is not hydrogen, Si(CH₃)₃, CH₂Si(CH₃)₃, methyl,        (CH₂)NH₂, (CH₂)₂NH₂, (CH₂)NHSO₂CH₃, or (CH₂)_(n)NHC(O)R^(1x); n        is 1 or 2; R^(1x) is methyl, C₂ alkene, cyclohexyl, cyclopentyl,        tetrahydrofuranyl, furanyl, or pyrrolidinyl, where the alkene,        cyclohexyl, cyclopentyl, tetrahydrofuranyl, furanyl, or        pyrrolidinyl is optionally substituted with one or two groups        independently selected from oxo and cyano;    -   wherein in Formula (A″), when X is CH, B is unsubstituted        phenyl, and W^(d) is

-   -    then R¹ is not phenyl;        or a pharmaceutically acceptable form thereof.

In certain embodiments, provided herein are compounds of Formula (I′) orFormula (A′):

wherein:R¹ is hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,aryl, heteroaryl, —COR², —COOR³, or —CONR⁴R⁵;B is hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,aryl, heteroaryl, —COR², —COOR³, —CONR⁴R⁵, or —Si(R⁶)₃;

-   -   wherein R², R³, R⁴, R⁵, and R⁶ are each, independently,        hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,        aryl, or heteroaryl;        W^(d) is heteroaryl, cycloalkyl, heterocycloalkyl, or aryl; and        X is CR^(1a) or N;    -   wherein R^(1a) is hydrogen, halo, alkyl, alkenyl, alkynyl, or        CN;    -   wherein each alkyl, alkenyl, or alkynyl is optionally        substituted with one or more halo, haloalkyl, OH, alkoxy, NH₂,        NH(alkyl), N(alkyl)₂, COH, CO(alkyl), COOH, COO(alkyl), CONH₂,        CONH(alkyl), CON(alkyl)₂, S(O)(alkyl), S(O)₂(alkyl), cycloalkyl,        heterocycloalkyl, aryl or heteroaryl;    -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, haloalkyl, alkyl,        alkenyl, alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂,        COH, CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl),        CON(alkyl)₂, S(O)(alkyl), or S(O)₂(alkyl);    -   wherein in Formula (I″), when X is CH, B is unsubstituted        phenyl, and W^(d) is

-   -    then R¹ is not hydrogen, Si(CH₃)₃, CH₂Si(CH₃)₃, methyl,        (CH₂)NH₂, (CH₂)₂NH₂, (CH₂)NHSO₂CH₃, or (CH₂)_(n)NHC(O)R^(1x); n        is 1 or 2; R^(1x) is methyl, C₂ alkene, cyclohexyl, cyclopentyl,        tetrahydrofuranyl, furanyl, or pyrrolidinyl, where the alkene,        cyclohexyl, cyclopentyl, tetrahydrofuranyl, furanyl, or        pyrrolidinyl is optionally substituted with one or two groups        independently selected from oxo and cyano;    -   wherein in Formula (A″), when X is CH, B is unsubstituted        phenyl, and W^(d) is

-   -    then R¹ is not phenyl;        or a pharmaceutically acceptable form thereof.

In certain embodiments, provided herein are compounds of Formula (I) orFormula (A):

wherein:R¹ is hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,aryl, heteroaryl, —COR², —COOR³, or —CONR⁴R⁵;B is hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,aryl, heteroaryl, —COR², —COOR³, —CONR⁴R⁵, or —Si(R⁶)₃;

-   -   wherein R², R³, R⁴, R⁵, and R⁶ are each, independently,        hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,        aryl, or heteroaryl;        W^(d) is heteroaryl, cycloalkyl, heterocycloalkyl, or aryl; and        X is CR^(1a) or N;    -   wherein R^(1a) is hydrogen, halo, alkyl, alkenyl, alkynyl, or        CN;    -   wherein each alkyl, alkenyl, or alkynyl is optionally        substituted with one or more halo, OH, alkoxy, NH₂, NH(alkyl),        N(alkyl)₂, COH, CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl),        CON(alkyl)₂, S(O)(alkyl), S(O)₂(alkyl), cycloalkyl,        heterocycloalkyl, aryl or heteroaryl;    -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, alkyl, alkenyl,        alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂, COH,        CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl), CON(alkyl)₂,        S(O)(alkyl), or S(O)₂(alkyl);    -   wherein in Formula (I″), when X is CH, B is unsubstituted        phenyl, and W^(d) is

-   -    then R¹ is not hydrogen, Si(CH₃)₃, CH₂Si(CH₃)₃, methyl,        (CH₂)NH₂, (CH₂)₂NH₂, (CH₂)NHSO₂CH₃, or (CH₂)^(n)NHC(O)R^(1x); n        is 1 or 2; R^(1x) is methyl, C₂ alkene, cyclohexyl, cyclopentyl,        tetrahydrofuranyl, furanyl, or pyrrolidinyl, where the alkene,        cyclohexyl, cyclopentyl, tetrahydrofuranyl, furanyl, or        pyrrolidinyl is optionally substituted with one or two groups        independently selected from oxo and cyano;    -   wherein in Formula (A″), when X is CH, B is unsubstituted        phenyl, and W^(d) is

-   -    then R¹ is not phenyl;        or a pharmaceutically acceptable form thereof.

In certain embodiments, provided herein is a mixture of compounds ofFormula (I″), (I′), (I), (A″), (A′), or (A) wherein individual compoundsof the mixture exist predominately in an (S)- or (R)-isomericconfiguration. For example, the compound mixture has an (S)-enantiomericpurity of greater than about 55%, about 60%, about 65%, about 70%, about75%, about 80%, about 85%, about 90%, about 95%, about 96%, about 97%,about 98%, about 99%, about 99.5%, or more. In other embodiments, thecompound mixture has an (S)-enantiomeric purity of greater than about55% to about 99.5%, greater than about about 60% to about 99.5%, greaterthan about 65% to about 99.5%, greater than about 70% to about 99.5%,greater than about 75% to about 99.5%, greater than about 80% to about99.5%, greater than about 85% to about 99.5%, greater than about 90% toabout 99.5%, greater than about 95% to about 99.5%, greater than about96% to about 99.5%, greater than about 97% to about 99.5%, greater thanabout 98% to greater than about 99.5%, greater than about 99% to about99.5%, or more.

In other embodiments, the compound mixture has an (R)-enantiomericpurity of greater than about 55%, about 60%, about 65%, about 70%, about75%, about 80%, about 85%, about 90%, about 95%, about 96%, about 97%,about 98%, about 99%, about 99.5%, or more. In some other embodiments,the compound mixture has an (R)-enantiomeric purity of greater thanabout 55% to about 99.5%, greater than about about 60% to about 99.5%,greater than about 65% to about 99.5%, greater than about 70% to about99.5%, greater than about 75% to about 99.5%, greater than about 80% toabout 99.5%, greater than about 85% to about 99.5%, greater than about90% to about 99.5%, greater than about 95% to about 99.5%, greater thanabout 96% to about 99.5%, greater than about 97% to about 99.5%, greaterthan about 98% to greater than about 99.5%, greater than about 99% toabout 99.5%, or more.

In certain embodiments, provided herein are compounds of Formula (I′):

or a pharmaceutically acceptable form thereof, wherein R¹, B, W^(d) andX are as defined herein.

In certain embodiments, provided herein are compounds of Formula (I):

or a pharmaceutically acceptable form thereof, wherein R¹, B, W^(d) andX are as defined herein.

In certain embodiments, R¹ is branched alkyl, 5- or 6-membered aryl, 5-or 6-membered heteroaryl, 5- or 6-membered cycloalkyl, or 5- or6-membered heterocycloalkyl,

cyclopropyl, or methyl,wherein R^(A) is OH, alkoxy, cycloalkyl, heterocycloalkyl, aryl, orheteroaryl;x is 1, 2, 3, 4, 5, or 6;R⁷, R⁸, and R⁹ are each, independently, hydrogen, OH, alkoxy, NH₂,NH(alkyl), N(alkyl)₂, alkyl, alkenyl, alkynyl, cycloalkyl,heterocycloalkyl, aryl, or heteroaryl.

In some embodiments, R^(A) is hydroxyl, alkoxy or heterocycloalkyl. Insome embodiments, R⁷, R⁸, and R⁹ are, independently, alkyl of 1-4carbons, amino, hydroxyl, or alkoxy of 1-4 carbons.

In certain embodiments, R¹ is a 5- to 10-membered heteroaryl. In certainembodiments, R¹ is a 5- or 6-membered heteroaryl. In certainembodiments, R¹ is a 6-membered heteroaryl. In certain embodiments, R¹is a pyridinyl. In certain embodiments, R¹ is a pyrimidinyl. In certainembodiments, R¹ is a 5-membered heteroaryl. In certain embodiments, R¹is a thiazolyl. In certain embodiments, R¹ is a pyrazolyl. In certainembodiments, R¹ is an imidazolyl. In certain embodiments, the heteroarylis substituted with one or more alkyl.

In some embodiments, R¹ is: methyl.

In some embodiments, B is phenyl substituted with 0, 1, 2, or 3occurrence(s) of R^(Z). In some embodiments, B is unsubstituted phenyl.In some embodiments, B is phenyl substituted with 1 or 2 occurrence(s)of R^(Z). In some embodiments, B is phenyl optionally substituted at thepara position with R^(z). In some embodiments, B is phenyl optionallymono-substituted at the meta position with R^(z). In some embodiments, Bis phenyl optionally mono-substituted at the ortho position with R^(z).In some embodiments, B is phenyl optionally di-substituted at the metapositions with R^(z). In some embodiments, B is phenyl optionallydi-substituted at the ortho positions with R^(z). In some embodiments, Bis phenyl optionally di-substituted at the meta and ortho positions withR^(z). In some embodiments, B is phenyl optionally di-substituted at themeta and para positions with R^(z). In some embodiments, B is phenyloptionally di-substituted at the ortho and para positions with R^(z). Insome embodiments, B is phenyl not substituted at the ortho positions. Insome embodiments, R^(Z) is halo or alkyl. In some embodiments, B ismethyl, isopropyl, or cyclopropyl. In some embodiments, B is cyclohexylor optionally substituted alkyl. In some embodiments, B is aryl,heteroaryl, cycloalkyl, or heterocycloalkyl. In some embodiments, B is5- or 6-membered aryl or 3- to 6-membered cycloalkyl. In someembodiments, B is

In some embodiments, B is one of the following moieties:

In some embodiments, B is selected from the moieties presented in Table1.

TABLE 1 Illustrative B moieties of the compounds described herein. Sub-class # B B-1

B-2

B-3 —CH(CH₃)₂ B-4

B-5

B-6

B-7

B-8

B-9

B-10

B-11

B-12

B-13

B-14

B-15

B-16

B-17

B-18

B-19

B-20

B-21

B-22

B-23

B-24

B-25

B-26

B-27

B-28

B-29

B-30

B-31

B-32

B-33

B-34

B-35

B-36

B-37

B-38

B-39

B-40

B-41

B-42

B-43

B-44

B-45

B-46

B-47

B-48

B-49

B-50

B-51

B-52

B-53

B-54

B-55

B-56

B-57

B-58

B-59

B-60

B-61

B-62

B-63

B-64

B-65

B-66

B-67

B-68

B-69

B-70

B-71

B-72

B-73

B-74

B-75

B-76

B-77

B-78

B-79

B-80

B-81

B-82

B-83

B-84

B-85

B-86

B-87 —CH₃ B-88 —CH₂CH₃ B-89

B-90

B-91

B-92

B-93

B-94

B-95

B-96

B-97

B-98

B-99

B-100

B-101

B-102

B-103

B-104

B-105

B106

B107

B108

B109

In some embodiments, W^(d) is aryl (e.g., a monocyclic aryl or abicyclic aryl). In some embodiments, W^(d) is substituted orunsubstituted phenyl. In some embodiments, W^(d) is bicyclic aryl (e.g.,substituted or unsubstituted naphthyl). In some embodiments, W^(d) is

In certain embodiments, W^(d) is heteroaryl (e.g., monocyclicheteroaryl, e.g., a monocyclic 5- or 6-membered heteroaryl; or bicyclicheteroaryl, e.g., a 5/6-bicyclic heteroaryl or a 6/6-bicyclicheteroaryl).

In some embodiments, W^(d) is

wherein

-   -   X₁, X₂ and X₃ are each independently C, CR¹³, or N;    -   X₄, X₅ and X₆ are each independently N, NR¹², CR¹³, S, or O; and    -   wherein each of the W_(d) group is optionally substituted with        one or more of R¹⁰, R¹¹, R¹², and R¹³, where R¹⁰, R¹¹, R¹² an        R¹³ are are each independently hydrogen, alkyl, heteroalkyl,        alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, arylalkyl,        heteroaryl, heteroarylalkyl, alkoxy, heterocyclyloxy, amido,        amino, acyl, acyloxy, alkoxycarbonyl, sulfonamido, halo, cyano,        hydroxyl, nitro, phosphate, urea, carbonate, or NR′R″ wherein R′        and R″ together with the nitrogen to which they are attached        form a cyclic moiety; and    -   the point of attachment is at any open position on the W_(d)        group.

In some embodiments, W^(d) is

wherein

-   -   X₁, X₂ and X₃ are each independently C, CR¹³, or N;    -   X₄, X₅ and X₆ are each independently N, NR¹², CR¹³, S, or O; and    -   R¹⁰, R¹¹, R¹², and R¹³ are each independently hydrogen, alkyl,        heteroalkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl,        arylalkyl, heteroaryl, heteroarylalkyl, alkoxy, heterocyclyloxy,        amido, amino, acyl, acyloxy, alkoxycarbonyl, sulfonamido, halo,        cyano, hydroxyl, nitro, phosphate, urea, carbonate, or NR′R″        wherein R′ and R″ together with the nitrogen to which they are        attached form a cyclic moiety.

In certain embodiments, X₁ is N. In some embodiments, X₁ is CR¹³. Insome embodiments, X₁ is C.

In certain embodiments, X₂ is N. In some embodiments, X₂ is CR¹³. Insome embodiments, X₂ is C.

In certain embodiments, X₃ is N. In some embodiments, X₃ is CR¹³.

In certain embodiments, X₄ is N. In some embodiments, X₄ is CR¹³. Insome embodiments, X₄ is S.

In certain embodiments, X₅ is NR¹². In some embodiments, X₅ is CR¹³. Insome embodiments, X₅ is O. In some embodiments, X₅ is S.

In certain embodiments, X₆ is N. In some embodiments, X₆ is NH. In someembodiments, X₆ is CR¹³. In some embodiments, X₆ is NH. In someembodiments, X₆ is O.

In some embodiments, each R¹⁰ is independently hydrogen, halo (e.g.,fluoro, chloro, or bromo), cyano, hydroxyl, alkyl (e.g., methyl or CF₃),alkoxyl, amino (e.g., cycloalkylamino (e.g., cyclopropylamino),alkylamino (e.g., methylamino or dimethylamino), or NH₂), aryl (e.g.,substituted or unsubstituted phenyl), heteroaryl (e.g., a 5- or6-membered heteroaryl, e.g., pyrazolyl, pyridinyl, among others),heterocyclyl (e.g., N-morpholinyl), or amido. In some embodiments, eachR¹⁰ is independently hydrogen, alkyl (e.g., methyl), amino (e.g.,cyclopropylamino, methylamino or NH₂), heterocyclyl (e.g.,N-morpholinyl), heteroaryl (e.g., 4-pyrazolyl), amido or halo (e.g.,chloro). In one embodiment, R¹⁰ is NH₂. In one embodiment, R¹⁰ is H.

In certain embodiments, each R¹¹ is independently hydrogen, halo (e.g.,fluoro, chloro, or bromo), cyano, hydroxyl, alkyl (e.g., methyl or CF₃),alkoxyl, amino (e.g., cycloalkylamino (e.g., cyclopropylamino),alkylamino (e.g., methylamino or dimethylamino), or NH₂), aryl (e.g.,substituted or unsubstituted phenyl), heteroaryl (e.g., a 5- or6-membered heteroaryl, e.g., pyrazolyl, pyridinyl, among others),heterocyclyl (e.g., N-morpholinyl), or amido. In some embodiments, eachR¹¹ is independently hydrogen, amino, halo (e.g., bromo), aryl (e.g.,phenyl) or alkyl (e.g., methyl). In one embodiment, R¹¹ is H.

In certain embodiments, each R¹² is independently hydrogen, halo (e.g.,fluoro, chloro, or bromo), cyano, hydroxyl, alkyl (e.g., methyl or CF₃),alkoxyl, amino (e.g., cycloalkylamino (e.g., cyclopropylamino),alkylamino (e.g., methylamino or dimethylamino), or NH₂), aryl (e.g.,substituted or unsubstituted phenyl), heteroaryl (e.g., a 5- or6-membered heteroaryl, e.g., pyrazolyl, pyridinyl, among others),heterocyclyl (e.g., N-morpholinyl), or amido. In some embodiments, eachR¹² is independently hydrogen, amino, or alkyl (e.g., methyl). In oneembodiment, R¹² is H.

In certain embodiments, each R¹³ is independently hydrogen, halo (e.g.,fluoro, chloro, or bromo), cyano, hydroxyl, alkyl (e.g., methyl or CF₃),alkoxyl, amino (e.g., cycloalkylamino (e.g., cyclopropylamino),alkylamino (e.g., methylamino or dimethylamino), or NH₂), aryl (e.g.,substituted or unsubstituted phenyl), heteroaryl (e.g., a 5- or6-membered heteroaryl, e.g., pyrazolyl, pyridinyl, among others),heterocyclyl (e.g., N-morpholinyl), or amido. In some embodiments, eachR¹³ is independently hydrogen, amino (e.g., NH₂), amido (e.g.,NH—C(═O)Me), or alkyl (e.g., methyl). In one embodiment, R¹³ is H.

In some embodiments, W^(d) is:

wherein one of X₁ and X₂ is C and the other is N; and R¹⁰, R¹¹, R¹², andR¹³ are as defined herein. In some embodiments, R¹⁰ is hydrogen, halo(e.g., fluoro, chloro, or bromo), cyano, hydroxyl, alkyl (e.g., methylor CF₃), alkoxyl, amino (e.g., cycloalkylamino (e.g., cyclopropylamino),alkylamino (e.g., methylamino or dimethylamino), or NH₂), aryl (e.g.,substituted or unsubstituted phenyl), heteroaryl (e.g., a 5- or6-membered heteroaryl, e.g., pyrazolyl, pyridinyl, among others),heterocyclyl (e.g., N-morpholinyl), or amido. In some embodiments, R¹⁰is hydrogen, alkyl (e.g., methyl), amino (e.g., cyclopropylamino,methylamino or NH₂), heterocyclyl (e.g., N-morpholinyl), heteroaryl(e.g., 4-pyrazolyl), amido or halo (e.g., chloro). In one embodiment,R¹⁰ is NH₂. In one embodiment, R¹⁰ is H. In specific embodiment, one ofX₁ and X₂ is C and the other is N; R¹⁰ is H or NH₂; and R¹¹, R¹², andR¹³ are as defined herein. In specific embodiments, all of R¹¹, R¹² areH. In specific embodiments, two of R¹¹, R¹², and R¹³ are H, and one ofR¹¹, R¹², and R¹³ is alkyl (e.g., methyl or CF₃), halo, cyano, aryl(e.g., phenyl), or heteroaryl (e.g., a 5- or 6-membered heteroaryl, suchas, pyridinyl, pyrimidinyl, pyrazolyl, thiazolyl, imidazolyl, amongothers); and in some embodiments, the aryl and heteroaryl is optionallysubstituted with one or more substituents, such as, for example, halo(e.g., F or CO, cyano, hydroxyl, alkyl (e.g., methyl or CF₃), alkoxyl(e.g., methoxy, OCF₃, ethoxy, or isopropyloxy), sulfonyl (e.g.,S(O)₂Me), sulfonamidyl (e.g., S(O)₂NH₂, S(O)₂NHMe, S(O)₂N(Me)₂,S(O)₂NH-i-Pr, S(O)₂NH-t-Bu, S(O)₂NH-c-Pr, S(O)₂NHPh,S(O)₂—N-pyrrolidinyl, S(O)₂—N-morpholinyl, S(O)₂—N-piperazinyl,S(O)₂-4-methyl-N-piperazinyl, NHS(O)₂Me, NHS(O)₂Et, NHS(O)₂-c-Pr), orsulfonylurea (e.g., NHS(O)₂N(Me)₂).

In some embodiments, W^(d) is:

wherein X₃ is N or CR¹³; and R¹⁰, R¹¹, R¹² and R¹³ are as definedherein. In specific embodiments, X₃ is N or CR¹³; R¹⁰ is H or NH₂; andR¹¹, R¹², and R¹³ are as defined herein. In specific embodiments, R¹⁰ isNH₂. In specific embodiments, X₃ is N. In specific embodiments, one ofR¹¹ and R¹² is H, and the other is alkyl (e.g., methyl or CF₃), halo,cyano, aryl (e.g., phenyl), or heteroaryl (e.g., a 5- or 6-memberedheteroaryl, such as, pyridinyl, pyrimidinyl, pyrazolyl, thiazolyl,imidazolyl, among others); and in some embodiments, the aryl andheteroaryl is optionally substituted with one or more substituents, suchas, for example, halo (e.g., F or Cl), cyano, hydroxyl, alkyl (e.g.,methyl or CF₃), alkoxyl (e.g., methoxy, OCF₃, ethoxy, or isopropyloxy),sulfonyl (e.g., S(O)₂Me), sulfonamidyl (e.g., S(O)₂NH₂, S(O)₂NHMe,S(O)₂N(Me)₂, S(O)₂NH-i-Pr, S(O)₂NH-t-Bu, S(O)₂NH-c-Pr, S(O)₂NHPh,S(O)₂—N-pyrrolidinyl, S(O)₂—N-morpholinyl, S(O)₂—N-piperazinyl,S(O)₂-4-methyl-N-piperazinyl, NHS(O)₂Me, NHS(O)₂-c-Pr), or sulfonylurea(e.g., NHS(O)₂N(Me)₂).

In some embodiments, W^(d) is:

wherein one of X₁ and X₂ is N and the other is CR¹³; and R¹⁰, R¹¹, R¹²,and R¹³ are as defined herein. In specific embodiment, one of X₁ and X₂is N and the other is CR¹³; R¹⁰ is H or NH₂; and R¹¹, R¹², and R¹³ areas defined herein. In specific embodiments, X₁ is N and X₂ is CR¹³. Inspecific embodiments, X₁ is N and X₂ is CH. In specific embodiments, R¹⁰is NH₂. In specific embodiments, R¹¹, R¹² and R¹³ are H. In specificembodiments, at least one of R¹¹, R¹² and R¹³ is not H. In specificembodiments, one occurrence of R¹¹, R¹² and R¹³ is not H and the otheroccurrences of R¹¹, R¹² and R¹³ are H, and the one occurrence of R¹¹,R¹² and R¹³ (which is not hydrogen) is alkyl (e.g., methyl or CF₃),halo, cyano, aryl (e.g., phenyl), or heteroaryl (e.g., a 5- or6-membered heteroaryl, such as, pyridinyl, pyrimidinyl, pyrazolyl,thiazolyl, imidazolyl, among others); and in some embodiments, the aryland heteroaryl is optionally substituted with one or more substituents,such as, for example, halo (e.g., F or Cl), cyano, hydroxyl, alkyl(e.g., methyl or CF₃), alkoxyl (e.g., methoxy, OCF₃, ethoxy, orisopropyloxy), sulfonyl (e.g., S(O)₂Me), sulfonamidyl (e.g., S(O)₂NH₂,S(O)₂NHMe, S(O)₂N(Me)₂, S(O)₂NH-i-Pr, S(O)₂NH-t-Bu, S(O)₂NH-c-Pr,S(O)₂NHPh, S(O)₂—N-pyrrolidinyl, S(O)₂—N-morpholinyl,S(O)₂—N-piperazinyl, S(O)₂-4-methyl-N-piperazinyl, NHS(O)₂Me,NHS(O)₂-c-Pr), or sulfonylurea (e.g., NHS(O)₂N(Me)₂).

In exemplary embodiments, W^(d) is one of the following moieties:

wherein R¹¹ and R¹² are as defined herein.

In some embodiments, W^(d) is

In some embodiments, W^(d) is

In some embodiments, W^(d) is

In some embodiments, W^(d) is

In some embodiments, X is CH. In some embodiments, X is N.

In some embodiments, in Formula (I″), (I′), or (I), when X is CH, B isunsubstituted phenyl, and W^(d) is

then R¹ is not hydrogen, Si(CH₃)₃, CH₂Si(CH₃)₃, methyl, (CH₂)NH₂,(CH₂)₂NH₂, (CH₂)NHSO₂CH₃, or (CH₂)_(n)NHC(O)R^(1x); n is 1 or 2; R^(1x)is methyl, C₂ alkene, cyclohexyl, cyclopentyl, tetrahydrofuranyl,furanyl, or pyrrolidinyl, where the alkene, cyclohexyl, cyclopentyl,tetrahydrofuranyl, furanyl, or pyrrolidinyl is optionally substitutedwith one or two groups independently selected from oxo and cyano.

In some embodiments, in Formula (I″), (I′), or (I), when when X is CH, Bis unsubstituted phenyl, and W^(d) is

then R¹ is not (CH₂)_(n)NHC(O)R^(1x); n is 1; R^(1x) istetrahydrofuranyl or pyrrolidinyl, where the tetrahydrofuranyl orpyrrolidinyl is optionally substituted with oxo.

In some embodiments, in Formula (A″), (A′), or (A), when X is CH, B isunsubstituted phenyl, and W^(d) is

then R¹ is not phenyl.

In some embodiments, the compound is a compound of formula II:

wherein R¹, B, and X are as defined herein. In some embodiments, R¹ ishydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl,heteroaryl, —COR², —COOR³, or —CONR⁴R⁵;B is hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,aryl, heteroaryl, —COR², —COOR³, —CONR⁴R⁵, or —Si(R⁶)₃;

-   -   wherein R², R³, R⁴, R⁵, and R⁶ are each, independently,        hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,        aryl, or heteroaryl;        X is CR^(1a) or N;    -   wherein R^(1a) is hydrogen, halo, alkyl, alkenyl, alkynyl, or        CN;    -   wherein each alkyl, alkenyl, or alkynyl is optionally        substituted with one or more halo, OH, alkoxy, NH₂, NH(alkyl),        N(alkyl)₂, COH, CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl),        CON(alkyl)₂, S(O)(alkyl), S(O)₂(alkyl), cycloalkyl,        heterocycloalkyl, aryl or heteroaryl;    -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, alkyl, alkenyl,        alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂, COH,        CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl), CON(alkyl)₂,        S(O)(alkyl), or S(O)₂(alkyl); and    -   wherein when X is CH, and B is unsubstituted phenyl, then R¹ is        not hydrogen, Si(CH₃)₃, CH₂Si(CH₃)₃, methyl, (CH₂)NH₂,        (CH₂)₂NH₂, (CH₂)NHSO₂CH₃, or (CH₂)_(n)NHC(O)R^(1x); n is 1 or 2;        R^(1x) is methyl, C₂ alkene, cyclohexyl, cyclopentyl,        tetrahydrofuranyl, furanyl, or pyrrolidinyl, where the alkene,        cyclohexyl, cyclopentyl, tetrahydrofuranyl, furanyl, or        pyrrolidinyl is optionally substituted with one or two groups        independently selected from oxo and cyano.

In some embodiments, the compound is a compound of formula III:

wherein R¹ and B are defined herein. In some embodiments, R¹ ishydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl,heteroaryl, —COR², —COOR³, or —CONR⁴R⁵;B is hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,aryl, heteroaryl, —COR², —COOR³, —CONR⁴R⁵, or —Si(R⁶)₃;

-   -   wherein R², R³, R⁴, R⁵, and R⁶ are each, independently,        hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,        aryl, or heteroaryl;    -   wherein each alkyl, alkenyl, or alkynyl is optionally        substituted with one or more halo, OH, alkoxy, NH₂, NH(alkyl),        N(alkyl)₂, COH, CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl),        CON(alkyl)₂, S(O)(alkyl), S(O)₂(alkyl), cycloalkyl,        heterocycloalkyl, aryl or heteroaryl;    -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, alkyl, alkenyl,        alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂, COH,        CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl), CON(alkyl)₂,        S(O)(alkyl), or S(O)₂(alkyl); and    -   wherein when B is unsubstituted phenyl, then R¹ is not hydrogen,        Si(CH₃)₃, CH₂Si(CH₃)₃, methyl, (CH₂)NH₂, (CH₂)₂NH₂,        (CH₂)NHSO₂CH₃, or (CH₂)_(n)NHC(O)R^(1x); n is 1 or 2; R^(1x) is        methyl, C₂ alkene, cyclohexyl, cyclopentyl, tetrahydrofuranyl,        furanyl, or pyrrolidinyl, where the alkene, cyclohexyl,        cyclopentyl, tetrahydrofuranyl, furanyl, or pyrrolidinyl is        optionally substituted with one or two groups independently        selected from oxo and cyano.

In some embodiments, the compound is a compound of formula IV:

wherein R¹ and B are as defined herein. In some embodiments, R¹ ishydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl,heteroaryl, —COR², —COOR³, or —CONR⁴R⁵; B is hydrogen, alkyl, alkenyl,alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, —COR², —COOR³,—CONR⁴R⁵, or —Si(R⁶)₃;

-   -   wherein R², R³, R⁴, R⁵, and R⁶ are each, independently,        hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,        aryl, or heteroaryl;    -   wherein each alkyl, alkenyl, or alkynyl is optionally        substituted with one or more halo, OH, alkoxy, NH₂, NH(alkyl),        N(alkyl)₂, COH, CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl),        CON(alkyl)₂, S(O)(alkyl), S(O)₂(alkyl), cycloalkyl,        heterocycloalkyl, aryl or heteroaryl; and    -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, alkyl, alkenyl,        alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂, COH,        CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl), CON(alkyl)₂,        S(O)(alkyl), or S(O)₂(alkyl).

In some embodiments of formula II, III, and IV, B is phenyl substitutedwith 0, 1, 2, or 3 occurrence(s) of R^(Z). In some embodiments, B isunsubstituted phenyl. In some embodiments, B is phenyl substituted with1 or 2 occurrence(s) of R^(Z). In some embodiments, R^(Z) is halo oralkyl. In some embodiments, B is methyl, isopropyl, or cyclopropyl. Insome embodiments, B is cyclohexyl or optionally substituted alkyl. Insome embodiments, B is aryl, heteroaryl, cycloalkyl, orheterocycloalkyl. In some embodiments, B is 5- or 6-membered aryl or 3-to 6-membered cycloalkyl. In some embodiments, B is

In some embodiments, the compound is a compound of formula V:

wherein R¹ and X are as defined herein. In some embodiments, R¹ ishydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl,heteroaryl, —COR², —COOR³, or —CONR⁴R⁵;

-   -   wherein R², R³, R⁴, and R⁵ are each, independently, hydrogen,        alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, or        heteroaryl;        X is CR^(1a) or N;    -   wherein R^(1a) is hydrogen, halo, alkyl, alkenyl, alkynyl, or        CN;    -   wherein each alkyl, alkenyl, or alkynyl is optionally        substituted with one or more halo, OH, alkoxy, NH₂, NH(alkyl),        N(alkyl)₂, COH, CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl),        CON(alkyl)₂, S(O)(alkyl), S(O)₂(alkyl), cycloalkyl,        heterocycloalkyl, aryl or heteroaryl;    -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, alkyl, alkenyl,        alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂, COH,        CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl), CON(alkyl)₂,        S(O)(alkyl), or S(O)₂(alkyl); and    -   wherein when X is CH, then R¹ is not hydrogen, Si(CH₃)₃,        CH₂Si(CH₃)₃, methyl, (CH₂)NH₂, (CH₂)₂NH₂, (CH₂)NHSO₂CH₃, or        (CH₂)_(n)NHC(O)R^(1x); n is 1 or 2; R^(1x) is methyl, C₂ alkene,        cyclohexyl, cyclopentyl, tetrahydrofuranyl, furanyl, or        pyrrolidinyl, where the alkene, cyclohexyl, cyclopentyl,        tetrahydrofuranyl, furanyl, or pyrrolidinyl is optionally        substituted with one or two groups independently selected from        oxo and cyano.

In some embodiments, the compound is a compound of formula VI:

wherein R¹ is as defined herein. In some embodiments, R¹ is hydrogen,alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl,—COR², —COOR³, or —CONR⁴R⁵;

-   -   wherein R², R³, R⁴, and R⁵ are each, independently, hydrogen,        alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, or        heteroaryl;    -   wherein each alkyl, alkenyl, or alkynyl is optionally        substituted with one or more halo, OH, alkoxy, NH₂, NH(alkyl),        N(alkyl)₂, COH, CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl),        CON(alkyl)₂, S(O)(alkyl), S(O)₂(alkyl), cycloalkyl,        heterocycloalkyl, aryl or heteroaryl;    -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, alkyl, alkenyl,        alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂, COH,        CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl), CON(alkyl)₂,        S(O)(alkyl), or S(O)₂(alkyl); and    -   wherein R¹ is not hydrogen, Si(CH₃)₃, CH₂Si(CH₃)₃, methyl,        (CH₂)NH₂, (CH₂)₂NH₂, (CH₂)NHSO₂CH₃, or (CH₂)_(n)NHC(O)R^(1x); n        is 1 or 2; R^(1x) is methyl, C₂ alkene, cyclohexyl, cyclopentyl,        tetrahydrofuranyl, furanyl, or pyrrolidinyl, where the alkene,        cyclohexyl, cyclopentyl, tetrahydrofuranyl, furanyl, or        pyrrolidinyl is optionally substituted with one or two groups        independently selected from oxo and cyano;

In some embodiments, the compound is a compound of formula VII:

wherein R¹ is as defined herein. In some embodiments, R¹ is hydrogen,alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl,—COR², —COOR³, or —CONR⁴R⁵;

-   -   wherein R², R³, R⁴, and R⁵ are each, independently, hydrogen,        alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, or        heteroaryl;    -   wherein each alkyl, alkenyl, or alkynyl is optionally        substituted with one or more halo, OH, alkoxy, NH₂, NH(alkyl),        N(alkyl)₂, COH, CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl),        CON(alkyl)₂, S(O)(alkyl), S(O)₂(alkyl), cycloalkyl,        heterocycloalkyl, aryl or heteroaryl; and    -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, alkyl, alkenyl,        alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂, COH,        CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl), CON(alkyl)₂,        S(O)(alkyl), or S(O)₂(alkyl).

In some embodiments, the compound is a compound of formula VIII:

wherein X and R¹ are as defined herein. In some embodiments, R¹ ishydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl,heteroaryl, —COR², —COOR³, or —CONR⁴R⁵;

-   -   wherein R², R³, R⁴, and R⁵ are each, independently, hydrogen,        alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, or        heteroaryl;        X is CR^(1a) or N;    -   wherein R^(1a) is hydrogen, halo, alkyl, alkenyl, alkynyl, or        CN;    -   wherein each alkyl, alkenyl, or alkynyl is optionally        substituted with one or more halo, OH, alkoxy, NH₂, NH(alkyl),        N(alkyl)₂, COH, CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl),        CON(alkyl)₂, S(O)(alkyl), S(O)₂(alkyl), cycloalkyl,        heterocycloalkyl, aryl or heteroaryl;    -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, alkyl, alkenyl,        alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂, COH,        CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl), CON(alkyl)₂,        S(O)(alkyl), or S(O)₂(alkyl); and    -   wherein when X is CH and W^(d) is

-   -    then R¹ is not hydrogen, Si(CH₃)₃, CH₂Si(CH₃)₃, methyl,        (CH₂)NH₂, (CH₂)₂NH₂, (CH₂)NHSO₂CH₃, or (CH₂)_(n)NHC(O)R^(1x); n        is 1 or 2; R^(1x) is methyl, C₂ alkene, cyclohexyl, cyclopentyl,        tetrahydrofuranyl, furanyl, or pyrrolidinyl, where the alkene,        cyclohexyl, cyclopentyl, tetrahydrofuranyl, furanyl, or        pyrrolidinyl is optionally substituted with one or two groups        independently selected from oxo and cyano.

In some embodiments of formulas II-VIII, R¹ is branched alkyl, 5- or6-membered aryl, 5- or 6-membered heteroaryl, 5- or 6-memberedcycloalkyl, or 5- or 6-membered heterocycloalkyl,

cyclopropyl, or methyl,wherein R^(A) is OH, alkoxy, cycloalkyl, heterocycloalkyl, aryl, orheteroaryl;x is 1, 2, 3, 4, 5, or 6;R⁷, R⁸, and R⁹ are each, independently, hydrogen, OH, alkoxy, NH₂,NH(alkyl), N(alkyl)₂, alkyl, alkenyl, alkynyl, cycloalkyl,heterocycloalkyl, aryl, or heteroaryl.

In some embodiments, R^(A) is hydroxyl, alkoxy or heterocycloalkyl. Insome embodiments, R⁷, R⁸, and R⁹ are, independently, alkyl of 1-4carbons, amino, hydroxyl, or alkoxy of 1-4 carbons.

In certain embodiments, R¹ is a 5- to 10-membered heteroaryl. In certainembodiments, R¹ is a 5- to 6-membered heteroaryl. In certainembodiments, R¹ is a 6-membered heteroaryl. In certain embodiments, R¹is a pyridinyl. In certain embodiments, R¹ is a pyrimidinyl. In certainembodiments, R¹ is a 5-membered heteroaryl. In certain embodiments, R¹is a thiazolyl. In certain embodiments, R¹ is a pyrazolyl. In certainembodiments, R¹ is an imidazolyl. In certain embodiments, the heteroarylis substituted with one or more alkyl.

In some embodiments of formulas II-VIII, R¹ is: methyl,

In some embodiments, the compound is a compound of formula IX:

wherein R¹, B, and W^(d) are as defined herein. In some embodiments, R¹is hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,aryl, heteroaryl, —COR², —COOR³, or —CONR⁴R⁵; B is hydrogen, alkyl,alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, —COR²,—COOR³, —CONR⁴R⁵, or —Si(R⁶)₃;

-   -   wherein R², R³, R⁴, R⁵, and R⁶ are each, independently,        hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,        aryl, or heteroaryl;        W^(d) is heteroaryl, cycloalkyl, heterocycloalkyl, or aryl; and    -   wherein each alkyl, alkenyl, or alkynyl is optionally        substituted with one or more halo, OH, alkoxy, NH₂, NH(alkyl),        N(alkyl)₂, COH, CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl),        CON(alkyl)₂, S(O)(alkyl), S(O)₂(alkyl), cycloalkyl,        heterocycloalkyl, aryl or heteroaryl;    -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, alkyl, alkenyl,        alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂, COH,        CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl), CON(alkyl)₂,        S(O)(alkyl), S(O)₂(alkyl); and    -   wherein when B is unsubstituted phenyl and W^(d) is

-   -    then R¹ is not hydrogen, Si(CH₃)₃, CH₂Si(CH₃)₃, methyl,        (CH₂)NH₂, (CH₂)₂NH₂, (CH₂)NHSO₂CH₃, or (CH₂)_(n)NHC(O)R^(1x); n        is 1 or 2; R^(1x) is methyl, C₂ alkene, cyclohexyl, cyclopentyl,        tetrahydrofuranyl, furanyl, or pyrrolidinyl, where the alkene,        cyclohexyl, cyclopentyl, tetrahydrofuranyl, furanyl, or        pyrrolidinyl is optionally substituted with one or two groups        independently selected from oxo and cyano.

In some embodiments, the compound is a compound of formula X:

wherein R¹, B, and W^(d) are as defined herein. In some embodiments, R¹is hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,aryl, heteroaryl, —COR², —COOR³, or —CONR⁴R⁵; B is hydrogen, alkyl,alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, —COR²,—COOR³, —CONR⁴R⁵, or —Si(R⁶)₃;

-   -   wherein R², R³, R⁴, R⁵, and R⁶ are each, independently,        hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,        aryl, or heteroaryl;        W^(d) is heteroaryl, cycloalkyl, heterocycloalkyl, or aryl; and    -   wherein each alkyl, alkenyl, or alkynyl is optionally        substituted with one or more halo, OH, alkoxy, NH₂, NH(alkyl),        N(alkyl)₂, COH, CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl),        CON(alkyl)₂, S(O)(alkyl), S(O)₂(alkyl), cycloalkyl,        heterocycloalkyl, aryl or heteroaryl; and    -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, alkyl, alkenyl,        alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂, COH,        CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl), CON(alkyl)₂,        S(O)(alkyl), or S(O)₂(alkyl).

In some embodiments of formulas I-X, R¹ is

wherein R^(A) and R⁷-R⁹ are as defined herein.

In certain embodiments, R¹ is a 5- to 10-membered heteroaryl. In certainembodiments, R¹ is a 5- to 6-membered heteroaryl. In certainembodiments, R¹ is a 6-membered heteroaryl. In certain embodiments, R¹is a pyridinyl. In certain embodiments, R¹ is a pyrimidinyl. In certainembodiments, R¹ is a 5-membered heteroaryl. In certain embodiments, R¹is a thiazolyl. In certain embodiments, R¹ is a pyrazolyl. In certainembodiments, R¹ is an imidazolyl. In certain embodiments, the heteroarylis substituted with one or more alkyl.

In some embodiments, the compound is a compound of formula XI:

wherein R¹ and W^(d) are as defined herein. In some embodiments, R¹ ishydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl,heteroaryl, —COR², —COOR³, or —CONR⁴R⁵;

-   -   wherein R², R³, R⁴, and R⁵ are each, independently, hydrogen,        alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, or        heteroaryl;        W^(d) is heteroaryl, cycloalkyl, heterocycloalkyl, or aryl; and    -   wherein each alkyl, alkenyl, or alkynyl is optionally        substituted with one or more halo, OH, alkoxy, NH₂, NH(alkyl),        N(alkyl)₂, COH, CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl),        CON(alkyl)₂, S(O)(alkyl), S(O)₂(alkyl), cycloalkyl,        heterocycloalkyl, aryl or heteroaryl; and    -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, alkyl, alkenyl,        alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂, COH,        CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl), CON(alkyl)₂,        S(O)(alkyl), or S(O)₂(alkyl).

In some embodiments, the compound is a compound of formula XII:

wherein R¹ and W^(d) are as defined herein. In some embodiments, R¹ ishydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl,heteroaryl, —COR², —COOR³, or —CONR⁴R⁵;

-   -   wherein R², R³, R⁴, and R⁵ are each, independently, hydrogen,        alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, or        heteroaryl;        W^(d) is heteroaryl, cycloalkyl, heterocycloalkyl, or aryl; and    -   wherein each alkyl, alkenyl, or alkynyl is optionally        substituted with one or more halo, OH, alkoxy, NH₂, NH(alkyl),        N(alkyl)₂, COH, CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl),        CON(alkyl)₂, S(O)(alkyl), S(O)₂(alkyl), cycloalkyl,        heterocycloalkyl, aryl or heteroaryl;    -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, alkyl, alkenyl,        alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂, COH,        CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl), CON(alkyl)₂,        S(O)(alkyl), or S(O)₂(alkyl); and    -   wherein when W^(d) is

-   -    then R¹ is not hydrogen, Si(CH₃)₃, CH₂Si(CH₃)₃, methyl,        (CH₂)NH₂, (CH₂)₂NH₂, (CH₂)NHSO₂CH₃, or (CH₂)_(n)NHC(O)R^(1x); n        is 1 or 2; R^(1x) is methyl, C₂ alkene, cyclohexyl, cyclopentyl,        tetrahydrofuranyl, furanyl, or pyrrolidinyl, where the alkene,        cyclohexyl, cyclopentyl, tetrahydrofuranyl, furanyl, or        pyrrolidinyl is optionally substituted with one or two groups        independently selected from oxo and cyano

In some embodiments, the compound is a compound of formula XIII,

wherein B and W^(d) are as defined herein. In some embodiments, B ishydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl,heteroaryl, —COR², —COOR³, —CONR⁴R⁵, or —Si(R⁶)₃;

-   -   wherein R², R³, R⁴, R⁵, and R⁶ are each, independently,        hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,        aryl, or heteroaryl;        W^(d) is heteroaryl, cycloalkyl, heterocycloalkyl, or aryl; and    -   wherein each alkyl, alkenyl, or alkynyl is optionally        substituted with one or more halo, OH, alkoxy, NH₂, NH(alkyl),        N(alkyl)₂, COH, CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl),        CON(alkyl)₂, S(O)(alkyl), S(O)₂(alkyl), cycloalkyl,        heterocycloalkyl, aryl or heteroaryl; and    -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, alkyl, alkenyl,        alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂, COH,        CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl), CON(alkyl)₂,        S(O)(alkyl), or S(O)₂(alkyl).

In some embodiments, the compound is a compound of formula XIV:

wherein B and W^(d) are as defined herein. In some embodiments, B ishydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl,heteroaryl, —COR², —COOR³, —CONR⁴R⁵, or —Si(R⁶)₃;

-   -   wherein R², R³, R⁴, R⁵, and R⁶ are each, independently,        hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,        aryl, or heteroaryl;        W^(d) is heteroaryl, cycloalkyl, heterocycloalkyl, or aryl; and    -   wherein each alkyl, alkenyl, or alkynyl is optionally        substituted with one or more halo, OH, alkoxy, NH₂, NH(alkyl),        N(alkyl)₂, COH, CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl),        CON(alkyl)₂, S(O)(alkyl), S(O)₂(alkyl), cycloalkyl,        heterocycloalkyl, aryl or heteroaryl; and    -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, alkyl, alkenyl,        alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂, COH,        CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl), CON(alkyl)₂,        S(O)(alkyl), or S(O)₂(alkyl).

In some embodiments the compound is a compound of formula XV:

wherein W^(d) is as defined herein. In some embodiments, W^(d) iscycloalkyl, heterocycloalkyl, aryl, or heteroaryl; and

-   -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, alkyl, alkenyl,        alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂, COH,        CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl), or CON(alkyl)₂.

In some embodiments, the compound is a compound of formula XVI:

wherein W^(d) is defined herein. In some embodiments, W^(d) iscycloalkyl, heterocycloalkyl, aryl, or heteroaryl; and

-   -   wherein each cycloalkyl, heterocycloalkyl, aryl or heteroaryl is        optionally substituted with one or more halo, alkyl, alkenyl,        alkynyl, OH, alkoxy, oxo, NH₂, NH(alkyl), N(alkyl)₂, COH,        CO(alkyl), COOH, COO(alkyl), CONH₂, CONH(alkyl), or CON(alkyl)₂.

In some embodiments of formulas IX-XVI, W^(d) is

In some embodiments of formulas IX-XVI, W^(d) is

In some embodiments of formulas IX-XVI, W^(d) is

In some embodiments of formulas IX-XVI, W^(d) is

In some embodiments of formulas I-XII, R¹ is not hydrogen. In someembodiments of formulas I-XII, R¹ is not linear alkyl or hydrogen. Insome embodiments of formulas I-XII, R¹ is not linear C₁-C₃ alkyl orhydrogen. In some embodiments of formulas I-XII, R¹ is not methyl orhydrogen.

In certain embodiments, provided herein are compounds of Formula (A):

or a pharmaceutically acceptable form thereof, wherein R¹, B, W^(d) andX are as defined herein. In certain embodiments, R¹ is alkyl orheteroaryl. In certain embodiments, R¹ is heteroaryl. In certainembodiments, R¹ is alkyl. In certain embodiments, B is phenyl. Incertain embodiments, X is CH or N. In certain embodiments, X is CH.In certain embodiments, X is N. In certain embodiments, W^(d) is

In certain embodiments, the compound of formula (A″), (A′), or (A) is amixture of trans and cis (e.g., where R¹ is trans or cis). In certainembodiments, R¹ is trans. In certain embodiments, R¹ is cis. In certainembodiments, the percentage of trans to cis is about 50%, greater thanabout 50%, greater than about 55%, greater than about 60%, greater thanabout 65%, greater than about 70%, greater than about 75%, greater thanabout 80%, greater than about 85%, greater than about 90%, greater thanabout 95%, greater than about 96%, greater than about 97%, greater thanabout 98%, or greater than about 99%.

In one embodiment, provided herein is a compound of Formula XVII:

wherein:R¹ and B are each, independently, linear or branched alkyl, alkenyl,alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, —COR², —COOR³,—CONR⁴R⁵, or —Si(R⁶)₃;

-   -   wherein R², R³, R⁴, R⁵, and R⁶ are each, independently,        hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl,        aryl, or heteroaryl;        W^(d) is heteroaryl, cycloalkyl, heterocycloalkyl, or aryl; and        X is CH or N;    -   wherein when X is CH, B is unsubstituted phenyl, W^(d) is

-   -    and R¹ is a linear alkyl, then the linear alkyl contains at        least three consecutively bonded carbons;    -   wherein when X is CH, B is unsubstituted phenyl, and W^(d) is

-   -    then R¹ is not Si(CH₃)₃;        or a pharmaceutically acceptable form thereof.

In some embodiments of Formula XVII, R¹ is branched alkyl, 5- or6-membered aryl, 5- or 6-membered heteroaryl, 5- or 6-memberedcycloalkyl, or 5- to 6-membered heterocycloalkyl,

cyclopropyl, or methyl,wherein R^(A) is hydroxyl, alkoxy, cycloalkyl, heterocycloalkyl, aryl,or heteroaryl;x is 1, 2, 3, 4, 5 or 6;R⁷, R⁸, and R⁹ are each, independently, hydrogen, hydroxyl, alkoxy,amino, alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, orheteroaryl, wherein at least two of R⁷, R⁸, and R⁹ are not hydrogen.

In some embodiments of Formula XVII, R^(A) is hydroxyl, alkoxy orheterocycloalkyl.

In some embodiments of Formula XVII, R⁷, R⁸, and R⁹ are, independently,alkyl of 1-4 carbons, amino, hydroxyl, or alkoxy of 1-4 carbons.

In some embodiments of Formula XVII, R₁ is: methyl,

In some embodiments of Formula XVII, B is phenyl substituted with 0, 1,2, or 3 occurrence(s) of R^(z). In some embodiments, B is unsubstitutedphenyl. In some embodiments, B is phenyl substituted with 1 or 2occurrence(s) of R^(Z). In some embodiments, R^(Z) is halo or alkyl. Insome embodiments, B is methyl, isopropyl, or cyclopropyl. In someembodiments, B is cyclohexyl or optionally substituted alkyl.

In some embodiments of Formula XVII, B is aryl, heteroaryl, cycloalkyl,or heterocycloalkyl.

In some embodiments of Formula XVII, B is 5- or 6-membered aryl or3-6-membered cycloalkyl.

In some embodiments of Formula XVII, B is

In some embodiments of Formula XVII, W^(d) is

In some embodiments of Formula XVII, W^(d) is

In some embodiments of Formula XVII, W^(d) is

In some embodiments of Formula XVII, W^(d) is

In some embodiments of Formula XVII, X is CH. In some embodiments, X isN.

In some embodiments of Formula XVII, when X is CH, B is unsubstitutedphenyl; W^(d) is

and R¹ is a linear alkyl, the linear alkyl contains at least fourconsecutively bonded carbons.

In some embodiments of Formula XVII, the compounds have the followingformula:

In some embodiments of Formula XVII, the compound has the followingformula:

In some embodiments, R¹ is a heteroaryl, aryl, cycloalkyl,heterocycloalkyl. In certain embodiments, R¹ is a heteroaryl. In certainembodiments, R¹ is a heteroaryl substituted with a ionic or polar group.In some embodiments, an ionic or polar group is a the part of a compoundincreases the solubility, stability, or metabolism or reduce theoff-target effect of the compound. In some embodiments, provided hereinis a compound of the following formulae:

wherein X, B, and W^(d) are as defined herein;Het is heteroaryl; andR^(1b) is halo, haloalkyl, alkyl, alkenyl, alkynyl, OH, alkoxy, oxo,NH₂, NH(alkyl), N(alkyl)₂, COH, CO(alkyl), COOH, COO(alkyl), CONH₂,CONH(alkyl), CON(alkyl)₂, S(O)(alkyl), S(O)₂(alkyl), S(O)₂NH₂,S(O)₂NH(alkyl), or S(O)₂N(alkyl)₂.

In certain embodiments, alkyl is C₁-C₈ alkyl. In certain embodiments,alkyl is C₁-C₆ alkyl. In another embodiment, alkyl is C₁-C₃ alkyl. Incertain embodiments, alkenyl is C₂-C₈ alkenyl. In certain embodiments,alkenyl is C₂-C₆ alkenyl. In another embodiment, alkenyl is C₂-C₃alkenyl. In certain embodiments, alkynyl is C₂-C₈ alkynyl. In certainembodiments, alkynyl is C₂-C₆ alkynyl. In another embodiment, alkynyl isC₂-C₃ alkynyl.

In certain embodiments, cycloalkyl is C₃-C₈ cycloalkyl. In certainembodiments, cycloalkyl is C₃-C₆ cycloalkyl. In certain embodiments,cycloalkyl is C₃-C₄ cycloalkyl. In certain embodiments, heterocycloalkylis a 3 to 14 membered saturated or partially saturated cycle containingone or more heteroatoms selected from a group consisting of N, O, and S.In certain embodiments, heterocycloalkyl is 3 to 10 membered. In anotherembodiment, heterocycloalkyl is 3 to 6 membered. In another embodiment,heterocycloalkyl is 6 membered. In certain embodiments, aryl is a C₆-C₁₄aromatic cycle. In certain embodiments, aryl is C₆-C₁₀. In anotherembodiment, aryl is C₆. In certain embodiments, heteroaryl is a 5 to 14membered aromatic cycle containing one or more heteroatoms selected froma group consisting of N, O, and S. In certain embodiments, heteroaryl is5 to 10 membered. In another embodiment, heteroaryl is 5 to 6 membered.In another embodiment, heteroaryl is 6 membered.

In certain embodiments, the compound provided herein is not a compoundselected from:

In certain embodiments, the compound of Formula (I″), (I′), (I), (A″),(A′), or (A) is in an (S)-stereochemical configuration.

In certain embodiments, the compound of Formula (I″), (I′), (I), (A″),(A′), or (A) is the S-enantiomer having an enantiomeric purity greaterthan 75%.

In certain embodiments, the compound of Formula (I′) or (A′) is acompound in Table 3, Table 4, Table 5, Table 6, Table 7, Table 8, Table9, Table 10, Table 11, Table 12, Table 13, or Table 14, or apharmaceutically acceptable form thereof.

In certain embodiments, the compound of Formula (I′) or (A′) is acompound in Table 3, Table 4, Table 5, or Table 6 or a pharmaceuticallyacceptable form thereof.

In certain embodiments, the compound of Formula (I′) or (A′) is acompound in Table 3 or Table 4 or a pharmaceutically acceptable formthereof. In certain embodiments, the compound of Formula (I′) or (A′) isa compound in Table 3 or a pharmaceutically acceptable form thereof. Incertain embodiments, the compound of Formula (I′) or (A′) is a compoundin Table 5 or a pharmaceutically acceptable form thereof. In certainembodiments, the compound of Formula (I′) or (A′) is a compound in Table7 or a pharmaceutically acceptable form thereof. In certain embodiments,the compound of Formula (I′) or (A′) is a compound in Table 9 or apharmaceutically acceptable form thereof. In certain embodiments, thecompound of Formula (I′) or (A′) is a compound in Table 11 or apharmaceutically acceptable form thereof. In certain embodiments, thecompound of Formula (I′) or (A′) is a compound in Table 13 or apharmaceutically acceptable form thereof. In certain embodiments, thecompound of Formula (I′) or (A′) is a compound in Table 4 or apharmaceutically acceptable form thereof. In certain embodiments, thecompound of Formula (I′) or (A′) is a compound in Table 6 or apharmaceutically acceptable form thereof. In certain embodiments, thecompound of Formula (I′) or (A′) is a compound in Table 8 or apharmaceutically acceptable form thereof. In certain embodiments, thecompound of Formula (I′) or (A′) is a compound in Table 10 or apharmaceutically acceptable form thereof. In certain embodiments, thecompound of Formula (I′) or (A′) is a compound in Table 12 or apharmaceutically acceptable form thereof. In certain embodiments, thecompound of Formula (I′) or (A′) is a compound in Table 14 or apharmaceutically acceptable form thereof

TABLE 3

Compound 1

Compound 2

Compound 3

Compound 4

Compound 5

Compound 6

Compound 7

Compound 8

Compound 9

Compound 10

Compound 11

Compound 12

Compound 13

Compound 14

Compound 15

Compound 16

Compound 17

Compound 18

Compound 19

Compound 20

Compound 21

Compound 22

Compound 23

Compound 24

Compound 25

Compound 26

Compound 27

Compound 28

Compound 29

Compound 30

Compound 31

Compound 32

Compound 33

Compound 34

Compound 35

Compound 36

Compound 37

Compound 38

Compound 39

Compound 40

Compound 41

Compound 42

Compound 43

Compound 44

Compound 45

Compound 46

Compound 47

Compound 48

Compound 49

Compound 50

Compound 51

Compound 52

Compound 53

Compound 54

Compound 55

Compound 56

Compound 57

Compound 58

Compound 59

Compound 60

Compound 61

Compound 62

Compound 63

Compound 64

Compound 65

Compound 66

Compound 67

Compound 68

Compound 69

Compound 70

Compound 71

Compound 72

Compound 73

Compound 74

Compound 75

Compound 76

Compound 77

Compound 78

Compound 79

Compound 80

Compound 81

Compound 82

Compound 83

Compound 84

Compound 85

Compound 86

Compound 87

Compound 88

Compound 89

Compound 90

Compound 91

Compound 92

Compound 93

Compound 94

Compound 95

Compound 96

Compound 97

Compound 98

Compound 99

Compound 100

Compound 101

Compound 102

Compound 103

Compound 104

Compound 105

Compound 106

Compound 107

Compound 108

Compound 109

Compound 110

TABLE 4

Compound 1001

Compound 1002

Compound 1003

Compound 1004

Compound 1005

Compound 1006

Compound 1007

Compound 1008

Compound 1009

Compound 1010

Compound 1011

Compound 1012

Compound 1013

Compound 1014

Compound 1015

Compound 1016

Compound 1017

Compound 1018

Compound 1019

Compound 1020

Compound 1021

Compound 1022

Compound 1023

Compound 1024

Compound 1025

Compound 1026

Compound 1027

Compound 1028

Compound 1029

Compound 1030

Compound 1031

Compound 1032

Compound 1033

Compound 1034

Compound 1035

Compound 1036

Compound 1037

Compound 1038

Compound 1039

Compound 1040

Compound 1041

Compound 1042

Compound 1043

Compound 1044

Compound 1045

Compound 1046

Compound 1047

Compound 1048

Compound 1049

Compound 1050

Compound 1051

Compound 1052

Compound 1053

Compound 1054

Compound 1055

Compound 1056

Compound 1057

Compound 1058

Compound 1059

Compound 1060

Compound 1061

Compound 1062

Compound 1063

Compound 1064

Compound 1065

Compound 1066

Compound 1067

Compound 1068

Compound 1069

Compound 1070

Compound 1071

Compound 1072

Compound 1073

Compound 1074

Compound 1075

Compound 1076

Compound 1077

Compound 1078

Compound 1079

Compound 1080

Compound 1081

Compound 1082

Compound 1083

Compound 1084

Compound 1085

Compound 1086

Compound 1087

Compound 1088

Compound 1089

Compound 1090

Compound 1091

Compound 1092

Compound 1093

Compound 1094

Compound 1095

TABLE 5

Compound 2001

Compound 2002

Compound 2003

Compound 2004

Compound 2005

Compound 2006

Compound 2007

Compound 2008

Compound 2009

Compound 2010

Compound 2011

Compound 2012

Compound 2013

Compound 2014

Compound 2015

Compound 2016

Compound 2017

Compound 2018

Compound 2019

Compound 2020

Compound 2021

Compound 2022

Compound 2023

Compound 2024

Compound 2025

Compound 2026

Compound 2027

Compound 2028

Compound 2029

Compound 2030

Compound 2031

Compound 2032

Compound 2033

Compound 2034

Compound 2035

Compound 2036

Compound 2037

Compound 2038

Compound 2039

Compound 2040

Compound 2041

Compound 2042

Compound 2043

Compound 2044

Compound 2045

Compound 2046

Compound 2047

Compound 2048

Compound 2049

Compound 2050

Compound 2051

Compound 2052

Compound 2053

Compound 2054

Compound 2055

Compound 2056

Compound 2057

Compound 2058

Compound 2059

Compound 2060

Compound 2061

Compound 2062

Compound 2063

Compound 2064

Compound 2065

Compound 2066

Compound 2067

Compound 2068

Compound 2069

Compound 2070

Compound 2071

Compound 2072

Compound 2073

Compound 2074

Compound 2075

Compound 2076

Compound 2077

Compound 2078

Compound 2079

Compound 2080

Compound 2081

Compound 2082

Compound 2083

Compound 2084

Compound 2085

Compound 2086

Compound 2087

Compound 2088

Compound 2089

Compound 2090

Compound 2091

Compound 2092

Compound 2093

Compound 2094

Compound 2095

Compound 2096

Compound 2097

Compound 2098

Compound 2099

Compound 2100

Compound 2101

Compound 2102

Compound 2103

Compound 2104

Compound 2105

Compound 2106

Compound 2107

Compound 2108

Compound 2109

Compound 2110

TABLE 6

Compound 3001

Compound 3002

Compound 3003

Compound 3004

Compound 3005

Compound 3006

Compound 3007

Compound 3008

Compound 3009

Compound 3010

Compound 3011

Compound 3012

Compound 3013

Compound 3014

Compound 3015

Compound 3016

Compound 3017

Compound 3018

Compound 3019

Compound 3020

Compound 3021

Compound 3022

Compound 3023

Compound 3024

Compound 3025

Compound 3026

Compound 3027

Compound 3028

Compound 3029

Compound 3030

Compound 3031

Compound 3032

Compound 3033

Compound 3034

Compound 3035

Compound 3036

Compound 3037

Compound 3038

Compound 3039

Compound 3040

Compound 3045

Compound 3046

Compound 3047

Compound 3048

Compound 3049

Compound 3050

Compound 3051

Compound 3052

Compound 3053

Compound 3054

Compound 3055

Compound 3056

Compound 3057

Compound 3058

Compound 3059

Compound 3060

Compound 3061

Compound 3062

Compound 3063

Compound 3064

Compound 3065

Compound 3066

Compound 3067

Compound 3068

Compound 3069

Compound 3070

Compound 3071

Compound 3072

Compound 3073

Compound 3074

Compound 3075

Compound 3076

Compound 3077

Compound 3078

Compound 3079

Compound 3080

Compound 3081

Compound 3082

Compound 3083

Compound 3084

Compound 3085

Compound 3086

Compound 3087

Compound 3088

Compound 3089

Compound 3090

Compound 3091

Compound 3092

Compound 3093

Compound 3094

Compound 3095

TABLE 7

Compound 1′

Compound 2′

Compound 3′

Compound 4′

Compound 5′

Compound 6′

Compound 7′

Compound 8′

Compound 9′

Compound 10′

Compound 11′

Compound 12′

Compound 13′

Compound 14′

Compound 15′

Compound 16′

Compound 17′

Compound 18′

Compound 19′

Compound 20′

Compound 21′

Compound 22′

Compound 23′

Compound 24′

Compound 25′

Compound 26′

Compound 27′

Compound 28′

Compound 29′

Compound 30′

Compound 31′

Compound 32′

Compound 33′

Compound 34′

Compound 35′

Compound 36A′

Compound 37′

Compound 38′

Compound 39′

Compound 40′

Compound 41′

Compound 42′

Compound 43′

Compound 44′

Compound 45′

Compound 46′

Compound 47′

Compound 48′

Compound 49′

Compound 50′

Compound 52′

Compound 53′

Compound 54′

Compound 55′

Compound 56′

Compound 57′

Compound 58′

Compound 59′

Compound 60′

Compound 61′

Compound 62′

Compound 63′

Compound 64′

Compound 65′

Compound 66′

Compound 67′

Compound 68′

Compound 69′

Compound 70′

Compound 71′

Compound 72′

Compound 73′

Compound 74′

Compound 75′

Compound 76′

Compound 77′

Compound 78′

Compound 79′

Compound 80′

Compound 81′

Compound 82′

Compound 83′

Compound 84′

Compound 85′

Compound 86′

Compound 87′

Compound 88′

Compound 89′

Compound 90′

Compound 91′

Compound 92′

Compound 93′

Compound 94′

Compound 95′

Compound 96′

Compound 97′

Compound 98′

Compound 99′

Compound 100′

Compound 101′

Compound 102′

Compound 103′

Compound 104′

Compound 105′

Compound 106′

Compound 108′

Compound 109′

Compound 110′

TABLE 8

Compound 1001′

Compound 1002′

Compound 1003′

Compound 1004′

Compound 1005′

Compound 1006′

Compound 1007′

Compound 1008′

Compound 1009′

Compound 1010′

Compound 1011′

Compound 1012′

Compound 1013′

Compound 1014′

Compound 1015′

Compound 1016′

Compound 1017′

Compound 1018′

Compound 1019′

Compound 1020′

Compound 1021′

Compound 1022′

Compound 1023′

Compound 1024′

Compound 1025′

Compound 1026′

Compound 1027′

Compound 1028′

Compound 1029′

Compound 1030′

Compound 1031′

Compound 1032′

Compound 1033′

Compound 1034′

Compound 1035′

Compound 1036′

Compound 1037′

Compound 1038′

Compound 1039′

Compound 1040′

Compound 1041′

Compound 1042′

Compound 1043′

Compound 1044′

Compound 1045′

Compound 1046′

Compound 1047′

Compound 1048′

Compound 1049′

Compound 1050′

Compound 1051′

Compound 1052′

Compound 1053′

Compound 1054′

Compound 1055′

Compound 1056′

Compound 1057′

Compound 1058′

Compound 1059′

Compound 1060′

Compound 1061′

Compound 1062′

Compound 1063′

Compound 1064′

Compound 1065′

Compound 1066′

Compound 1067′

Compound 1068′

Compound 1069′

Compound 1070′

Compound 1071′

Compound 1072′

Compound 1073′

Compound 1074′

Compound 1075′

Compound 1076′

Compound 1077′

Compound 1078′

Compound 1079′

Compound 1080′

Compound 1081′

Compound 1082′

Compound 1083′

Compound 1084′

Compound 1085′

Compound 1086′

Compound 1087′

Compound 1088′

Compound 1089′

Compound 1090′

Compound 1091′

Compound 1092′

Compound 1093′

Compound 1094′

Compound 1095′

TABLE 9

Compound 2001′

Compound 2002′

Compound 2003′

Compound 2004′

Compound 2005′

Compound 2006′

Compound 2007′

Compound 2008′

Compound 2009′

Compound 2010′

Compound 2011′

Compound 2012′

Compound 2013′

Compound 2014′

Compound 2015′

Compound 2016′

Compound 2017′

Compound 2018′

Compound 2019′

Compound 2020′

Compound 2021′

Compound 2022′

Compound 2023′

Compound 2024′

Compound 2025′

Compound 2026′

Compound 2027′

Compound 2028′

Compound 2029′

Compound 2030′

Compound 2031′

Compound 2032′

Compound 2033′

Compound 2034′

Compound 2035′

Compound 2036′

Compound 2037′

Compound 2038′

Compound 2039′

Compound 2040′

Compound 2041′

Compound 2042′

Compound 2043′

Compound 2044′

Compound 2045′

Compound 2046′

Compound 2047′

Compound 2048′

Compound 2049′

Compound 2050′

Compound 2052′

Compound 2053′

Compound 2054′

Compound 2055′

Compound 2056′

Compound 2057′

Compound 2058′

Compound 2059′

Compound 2060′

Compound 2061′

Compound 2062′

Compound 2063′

Compound 2064′

Compound 2065′

Compound 2066′

Compound 2067′

Compound 2068′

Compound 2069′

Compound 2070′

Compound 2071′

Compound 2072′

Compound 2073′

Compound 2074′

Compound 2075′

Compound 2076′

Compound 2077′

Compound 2078′

Compound 2079′

Compound 2080′

Compound 2081′

Compound 2082′

Compound 2083′

Compound 2084′

Compound 2085′

Compound 2086′

Compound 2087′

Compound 2088′

Compound 2089′

Compound 2090′

Compound 2091′

Compound 2092′

Compound 2093′

Compound 2094′

Compound 2095′

Compound 2096′

Compound 2097′

Compound 2098′

Compound 2099′

Compound 2100′

Compound 2101′

Compound 2102′

Compound 2103′

Compound 2104′

Compound 2105′

Compound 2106′

Compound 2108′

Compound 2109′

Compound 2110′

TABLE 10

Compound 3001′

Compound 3002′

Compound 3003′

Compound 3004′

Compound 3005′

Compound 3006′

Compound 3007′

Compound 3008′

Compound 3009′

Compound 3010′

Compound 3011′

Compound 3012′

Compound 3013′

Compound 3014′

Compound 3015′

Compound 3016′

Compound 3017′

Compound 3018′

Compound 3019′

Compound 3020′

Compound 3021′

Compound 3022′

Compound 3023′

Compound 3024′

Compound 3025′

Compound 3026′

Compound 3027′

Compound 3028′

Compound 3029′

Compound 3030′

Compound 3031′

Compound 3032′

Compound 3033′

Compound 3034′

Compound 3035′

Compound 3036′

Compound 3037′

Compound 3038′

Compound 3039′

Compound 3040′

Compound 3045′

Compound 3046′

Compound 3047′

Compound 3048′

Compound 3049′

Compound 3050′

Compound 3051′

Compound 3052′

Compound 3053′

Compound 3054′

Compound 3055′

Compound 3056′

Compound 3057′

Compound 3058′

Compound 3059′

Compound 3060′

Compound 3061′

Compound 3062′

Compound 3063′

Compound 3064′

Compound 3065′

Compound 3066′

Compound 3067′

Compound 3068′

Compound 3069′

Compound 3070′

Compound 3071′

Compound 3072′

Compound 3073′

Compound 3074′

Compound 3075′

Compound 3076′

Compound 3077′

Compound 3078′

Compound 3079′

Compound 3080′

Compound 3081′

Compound 3082′

Compound 3083′

Compound 3084′

Compound 3085′

Compound 3086′

Compound 3087′

Compound 3088′

Compound 3089′

Compound 3090′

Compound 3091′

Compound 3092′

Compound 3093′

Compound 3094′

Compound 3095′

TABLE 11

Compound 1r

Compound 2r

Compound 3r

Compound 4r

Compound 5r

Compound 6r

Compound 7r

Compound 8r

Compound 9r

Compound 10r

Compound 11r

Compound 12r

Compound 13r

Compound 14r

Compound 15r

Compound 16r

Compound 17r

Compound 18r

Compound 19r

Compound 20r

Compound 21r

Compound 22r

Compound 23r

Compound 24r

Compound 25r

Compound 26r

Compound 27r

Compound 28r

Compound 29r

Compound 30r

Compound 31r

Compound 32r

Compound 33r

Compound 34r

Compound 35r

Compound 36r

Compound 37r

Compound 38r

Compound 39r

Compound 40r

Compound 41r

Compound 42r

Compound 43r

Compound 44r

Compound 45r

Compound 46r

Compound 47r

Compound 48r

Compound 49r

Compound 50r

Compound 51r

Compound 52r

Compound 53r

Compound 54r

Compound 55r

Compound 56r

Compound 57r

Compound 58r

Compound 59r

Compound 60r

Compound 61r

Compound 62r

Compound 63r

Compound 64r

Compound 65r

Compound 66r

Compound 67r

Compound 68r

Compound 69r

Compound 70r

Compound 71r

Compound 72r

Compound 73r

Compound 74r

Compound 75r

Compound 76r

Compound 77r

Compound 78r

Compound 79r

Compound 80r

Compound 81r

Compound 82r

Compound 83r

Compound 84r

Compound 85r

Compound 86r

Compound 87r

Compound 88r

Compound 89r

Compound 90r

Compound 91r

Compound 92r

Compound 93r

Compound 94r

Compound 95r

Compound 96r

Compound 97r

Compound 98r

Compound 99r

Compound 100r

Compound 101r

Compound 102r

Compound 103r

Compound 104r

Compound 105r

Compound 106r

Compound 108r

Compound 109r

Compound 110r

TABLE 12

Compound 1001r

Compound 1002r

Compound 1003r

Compound 1004r

Compound 1005r

Compound 1006r

Compound 1007r

Compound 1008r

Compound 1009r

Compound 1010r

Compound 1011r

Compound 1012r

Compound 1013r

Compound 1014r

Compound 1015r

Compound 1016r

Compound 1017r

Compound 1018r

Compound 1019r

Compound 1020r

Compound 1021r

Compound 1022r

Compound 1023r

Compound 1024r

Compound 1025r

Compound 1026r

Compound 1027r

Compound 1028r

Compound 1029r

Compound 1030r

Compound 1031r

Compound 1032r

Compound 1033r

Compound 1034r

Compound 1035r

Compound 1036r

Compound 1037r

Compound 1038r

Compound 1039r

Compound 1040r

Compound 1041r

Compound 1042r

Compound 1043r

Compound 1044r

Compound 1045r

Compound 1046r

Compound 1047r

Compound 1048r

Compound 1049r

Compound 1050r

Compound 1051r

Compound 1052r

Compound 1053r

Compound 1054r

Compound 1055r

Compound 1056r

Compound 1057r

Compound 1058r

Compound 1059r

Compound 1060r

Compound 1061r

Compound 1062r

Compound 1063r

Compound 1064r

Compound 1065r

Compound 1066r

Compound 1067r

Compound 1068r

Compound 1069r

Compound 1070r

Compound 1071r

Compound 1072r

Compound 1073r

Compound 1074r

Compound 1075r

Compound 1076r

Compound 1077r

Compound 1078r

Compound 1079r

Compound 1080r

Compound 1081r

Compound 1082r

Compound 1083r

Compound 1084r

Compound 1085r

Compound 1086r

Compound 1087r

Compound 1088r

Compound 1089r

Compound 1090r

Compound 1091r

Compound 1092r

Compound 1093r

Compound 1094r

Compound 1095r

TABLE 13

Compound 2001r

Compound 2002r

Compound 2003r

Compound 2004r

Compound 2005r

Compound 2006r

Compound 2007r

Compound 2008r

Compound 2009r

Compound 2010r

Compound 2011r

Compound 2012r

Compound 2013r

Compound 2014r

Compound 2015r

Compound 2016r

Compound 2017r

Compound 2018r

Compound 2019r

Compound 2020r

Compound 2021r

Compound 2022r

Compound 2023r

Compound 2024r

Compound 2025r

Compound 2026r

Compound 2027r

Compound 2028r

Compound 2029r

Compound 2030r

Compound 2031r

Compound 2032r

Compound 2033r

Compound 2034r

Compound 2035r

Compound 2036r

Compound 2037r

Compound 2038r

Compound 2039r

Compound 2040r

Compound 2041r

Compound 2042r

Compound 2043r

Compound 2044r

Compound 2045r

Compound 2046r

Compound 2047r

Compound 2048r

Compound 2049r

Compound 2050r

Compound 2051r

Compound 2052r

Compound 2053r

Compound 2054r

Compound 2055r

Compound 2056r

Compound 2057r

Compound 2058r

Compound 2059r

Compound 2060r

Compound 2061r

Compound 2062r

Compound 2063r

Compound 2064r

Compound 2065r

Compound 2066r

Compound 2067r

Compound 2068r

Compound 2069r

Compound 2070r

Compound 2071r

Compound 2072r

Compound 2073r

Compound 2074r

Compound 2075r

Compound 2076r

Compound 2077r

Compound 2078r

Compound 2079r

Compound 2080r

Compound 2081r

Compound 2082r

Compound 2083r

Compound 2084r

Compound 2085r

Compound 2086r

Compound 2087r

Compound 2088r

Compound 2089r

Compound 2090r

Compound 2091r

Compound 2092r

Compound 2093r

Compound 2094r

Compound 2095r

Compound 2096r

Compound 2097r

Compound 2098r

Compound 2099r

Compound 2100r

Compound 2101r

Compound 2102r

Compound 2103r

Compound 2104r

Compound 2105r

Compound 2106r

Compound 2108r

Compound 2109r

Compound 2110r

TABLE 14

Compound 3001r

Compound 3002r

Compound 3003r

Compound 3004r

Compound 3005r

Compound 3006r

Compound 3007r

Compound 3008r

Compound 3009r

Compound 3010r

Compound 3011r

Compound 3012r

Compound 3013r

Compound 3014r

Compound 3015r

Compound 3016r

Compound 3017r

Compound 3018r

Compound 3019r

Compound 3020r

Compound 3021r

Compound 3022r

Compound 3023r

Compound 3024r

Compound 3025r

Compound 3026r

Compound 3027r

Compound 3028r

Compound 3029r

Compound 3030r

Compound 3031r

Compound 3032r

Compound 3033r

Compound 3034r

Compound 3035r

Compound 3036r

Compound 3037r

Compound 3038r

Compound 3039r

Compound 3040r

Compound 3045r

Compound 3046r

Compound 3047r

Compound 3048r

Compound 3049r

Compound 3050r

Compound 3051r

Compound 3052r

Compound 3053r

Compound 3054r

Compound 3055r

Compound 3056r

Compound 3057r

Compound 3058r

Compound 3059r

Compound 3060r

Compound 3061r

Compound 3062r

Compound 3063r

Compound 3064r

Compound 3065r

Compound 3066r

Compound 3067r

Compound 3068r

Compound 3069r

Compound 3070r

Compound 3071r

Compound 3072r

Compound 3073r

Compound 3074r

Compound 3075r

Compound 3076r

Compound 3077r

Compound 3078r

Compound 3079r

Compound 3080r

Compound 3081r

Compound 3082r

Compound 3083r

Compound 3084r

Compound 3085r

Compound 3086r

Compound 3087r

Compound 3088r

Compound 3089r

Compound 3090r

Compound 3091r

Compound 3092r

Compound 3093r

Compound 3094r

Compound 3095r

In some embodiments, the compound provided herein is:

or a pharmaceutically acceptable form thereof.

Provided herein, Compound AA has the structure:

Compound AA is disclosed as Compound 359 in International ApplicationWO2013/032591. As described in Table 4 of International ApplicationWO2013/032591, Compound AA has a PI3K-delta IC50 of less than 100 nM, aPI3K-gamma IC50 of 1 uM to 100 nM, a PI3K-alpha IC50 of greater than 1uM to 10 uM, and a PI3K-beta IC50 of greater than 1 uM to 10 uM.

Provided herein, compound BB is Compound 4 of the structure:

In some embodiments, one or more compounds described herein bind to aPI3 kinase (e.g., bind selectively). In some embodiments, one or morecompounds described herein bind selectively to a γ- or δ-subtype of aPI3 kinase. In some embodiments, one or more compounds described hereinbind selectively to a γ-subtype of a PI3 kinase. In some embodiments,one or more compounds described herein bind selectively to a δ-subtypeof a PI3 kinase. In one embodiment, one or more compounds describedherein selectively binds to δ over γ. In one embodiment, one or morecompounds described herein selectively binds to γ over δ.

In certain embodiments provided herein are methods of treating orpreventing a PI3K mediated disorder in a subject, the method comprisingadministering a therapeutically effective amount of a compound providedherein or composition provided herein to said subject. In certainembodiments, provided herein is the use of a compound provided herein inthe manufacture of a medicament for treating or preventing a PI3Kmediated disorder in a subject. In certain embodiments, a compoundprovided herein is for use in treating or preventing a PI3K mediateddisorder in a subject. In certain embodiments, the disorder is cancer,an inflammatory disease, or an auto-immune disease. In certainembodiments, the PI3K mediated disorder is a PI3K-γ mediated disorder.In certain embodiments, the PI3K mediated disorder is a PI3K-δ mediateddisorder. In certain embodiments, provided herein are methods forselectively inhibiting PI3K gamma over PI3K delta in a cell or subjectcomprising contacting the cell or administering to the subject acompound provided herein. In certain embodiments, provided wherein aremethods for selectively inhibiting PI3K gamma over PI3K delta in a cellor subject comprising contacting the cell or administering to thesubject

-   -   (i) a compound selected from compound 2, 4, 7, 9, 17, 19, 21,        26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73, 75, 77,        79, 80, 81, 88, and 89; or    -   (ii) a compound is selected from compound 1, 3, 6, 10, 11, 12,        16, 18, 20, 22, 25, 28, 34, 39, 42, 43, 53, 55, 59, 64, 65, 66,        67, 70, 76, 78, 82, 83, 84, 85, 86, and 90; or    -   (iii) a compound selected from compound 8, 13, 15, 23, 29, 33,        45, 51, 54, 57, and 68; or    -   (iv) a compound selected from compound 5, 14, 24, 31, 36, 46,        50, 69, 72, 74, and 91.

In certain embodiments, the compound is selected from compound 2, 4, 7,9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73,75, 77, 79, 80, 81, 88, and 89. In certain embodiments, the compound isselected from compound 1, 3, 6, 10, 11, 12, 16, 18, 20, 22, 25, 28, 34,39, 42, 43, 53, 55, 59, 64, 65, 66, 67, 70, 76, 78, 82, 83, 84, 85, 86,and 90. In certain embodiments, the compound is selected from compound8, 13, 15, 23, 29, 33, 45, 51, 54, 57, and 68. In certain embodiments,the compound is selected from compound 5, 14, 24, 31, 36, 46, 50, 69,72, 74, and 91.

In certain embodiments, provided herein are methods of synthesizing acompound provided herein. Provided herein are methods of making a PI3K-γselective compound comprising synthesizing a compound containing both(a) a non-terminal alkyne substituted bicyclic heterocyclic group and(b) an amido group. In some embodiments, the compound selectively bindsto PI3K-γ over PI3K-δ.

In some embodiments, the IC₅₀ of a compound provided herein for p110α,p110β, p110γ, or p110δ is less than about 1 μM, less than about 100 nM,less than about 50 nM, less than about 10 nM, less than 1 nM, or evenless than about 0.5 nM.

In some embodiments, non-limiting exemplary compounds exhibit one ormore functional characteristics disclosed herein. For example, one ormore compounds provided herein bind specifically to a PI3 kinase. Insome embodiments, the IC₅₀ of a compound provided herein for p110α,p110β, p110γ, or p110δ is less than about 1 μM, less than about 100 nM,less than about 50 nM, less than about 10 nM, less than about 1 nM, lessthan about 0.5 nM, less than about 100 pM, or less than about 50 pM.

In some embodiments, one or more of the compounds provided herein canselectively inhibit one or more members of type I or class Iphosphatidylinositol 3-kinases (PI3-kinase) with an IC₅₀ value of about100 nM, about 50 nM, about 10 nM, about 5 nM, about 100 pM, about 10 pM,or about 1 pM, or less, as measured in an in vitro kinase assay.

In some embodiments, one or more of the compounds provided herein canselectively inhibit one or two members of type I or class Iphosphatidylinositol 3-kinases (PI3-kinase), such as, PI3-kinase α,PI3-kinase β, PI3-kinase γ, and PI3-kinase δ. In some aspects, some ofthe compounds provided herein selectively inhibit PI3-kinase δ ascompared to all other type I PI3-kinases. In other aspects, some of thecompounds provided herein selectively inhibit PI3-kinase δ andPI3-kinase γ as compared to the rest of the type I PI3-kinases. In otheraspects, some of the compounds provided herein selectively inhibitPI3-kinase γ as compared to all other type I PI3-kinases.

In yet another aspect, an inhibitor that selectively inhibits one ormore members of type I PI3-kinases, or an inhibitor that selectivelyinhibits one or more type I PI3-kinase mediated signaling pathways,alternatively can be understood to refer to a compound that exhibits a50% inhibitory concentration (IC₅₀) with respect to a given type IPI3-kinase, that is at least about 10-fold, at least about 20-fold, atleast about 50-fold, at least about 100-fold, at least about 200-fold,at least about 500-fold, at least about 1000-fold, at least about2000-fold, at least about 5000-fold, or at least about 10,000-fold,lower than the inhibitor's IC₅₀ with respect to the rest of the othertype I PI3-kinases. In one embodiment, an inhibitor selectively inhibitsPI3-kinase δ as compared to PI3-kinase β with at least about 10-foldlower IC₅₀ for PI3-kinase δ. In certain embodiments, the IC₅₀ forPI3-kinase δ is below about 100 nM, while the IC₅₀ for PI3-kinase β isabove about 1000 nM. In certain embodiments, the IC₅₀ for PI3-kinase δis below about 50 nM, while the IC₅₀ for PI3-kinase β is above about5000 nM. In certain embodiments, the IC₅₀ for PI3-kinase δ is belowabout 10 nM, while the IC₅₀ for PI3-kinase β is above about 1000 nM,above about 5,000 nM, or above about 10,000 nM. In one embodiment, aninhibitor selectively inhibits PI3-kinase γ as compared to PI3-kinase βwith at least about 10-fold lower IC₅₀ for PI3-kinase γ. In certainembodiments, the IC₅₀ for PI3-kinase γ is below about 100 nM, while theIC₅₀ for PI3-kinase β is above about 1000 nM. In certain embodiments,the IC₅₀ for PI3-kinase γ is below about 50 nM, while the IC₅₀ forPI3-kinase β is above about 5000 nM. In certain embodiments, the IC₅₀for PI3-kinase γ is below about 10 nM, while the IC₅₀ for PI3-kinase βis above about 1000 nM, above about 5,000 nM, or above about 10,000 nM.

A PI3K gamma compound, e.g., a PI3K gamma inhibitor, can inhibit PI3Kgamma and can optionally also inhibit other PI3K isoforms such as PI3Kdelta. In one embodiment, a PI3K gamma inhibitor is also an inhibitor ofPI3K delta, i.e., a dual inhibitor of PI3K gamma and PI3K delta. In oneembodiment, the dual inhibitor has an IC50 for PI3K gamma that is within10-fold of its IC50 for PI3K delta.

A PI3K gamma selective compound, e.g., a PI3K gamma selective inhibitor,can exhibit a 50% inhibitory concentration (IC50) with respect to PI3Kgamma, that is at least about 10-fold lower than the compound's IC50with respect to the rest of the other type I PI3-kinases. In someembodiments, the PI3K gamma selective compound exhibits a 50% inhibitoryconcentration (IC50) with respect to PI3K gamma, that is at least aboutat least about 20-fold, at least about 30-fold, at least about 40-fold,at least about 50-fold, at least about 60-fold, at least about 70-fold,at least about 80-fold, at least about 90-fold, at least about 100-fold,at least about 200-fold, at least about 500-fold, at least about1000-fold, at least about 2000-fold, at least about 5000-fold, or atleast about 10,000-fold, lower than the compound's IC50 with respect tothe rest of the other type I PI3-kinases. In one embodiment, aninhibitor selectively inhibits PI3-kinase γ as compared to PI3-kinase δwith at least about 10-fold lower IC50 for PI3-kinase γ. In someembodiments, a PI3K gamma selective compound has an IC50 of about 1.8 nMfor PI3K gamma, a PI3K beta or PI3K delta IC50 of about 180 nM, an IC90value for PI3K gamma of about 16 nM, and an IC20 value for PI3K beta orPI3K delta of about 45 nM.

A PI3K delta compound, e.g., a PI3K delta inhibitor, can inhibit PI3Kdelta and can optionally also inhibit other PI3K isoforms such as PI3Kgamma. In one embodiment, a PI3K delta inhibitor is also an inhibitor ofPI3K gamma.

A PI3K delta selective compound, e.g., a PI3K delta selective inhibitor,can exhibit a 50% inhibitory concentration (IC50) with respect to PI3Kdelta, that is at least about 10-fold lower than the compound's IC50with respect to the rest of the other type I PI3-kinases. In someembodiments, the PI3K delta selective compound exhibits a 50% inhibitoryconcentration (IC50) with respect to PI3K delta, that is at least about20-fold, at least about 30-fold, at least about 40-fold, at least about50-fold, at least about 60-fold, at least about 70-fold, at least about80-fold, at least about 90-fold, at least about 100-fold, at least about200-fold, at least about 500-fold, at least about 1000-fold, at leastabout 2000-fold, at least about 5000-fold, or at least about10,000-fold, lower than the compound's IC50 with respect to the rest ofthe other type I PI3-kinases. In one embodiment, an inhibitorselectively inhibits PI3-kinase δ as compared to PI3-kinase γ with atleast about 10-fold lower IC50 for PI3-kinase δ.

Pharmaceutical Compositions

In some embodiments, provided herein are pharmaceutical compositionscomprising a compound as disclosed herein, or an enantiomer, a mixtureof enantiomers, or a mixture of two or more diastereomers thereof, or apharmaceutically acceptable form thereof (e.g., pharmaceuticallyacceptable salts, hydrates, solvates, isomers, prodrugs, andisotopically labeled derivatives), and a pharmaceutically acceptableexcipient, diluent, or carrier, including inert solid diluents andfillers, sterile aqueous solution and various organic solvents,permeation enhancers, solubilizers and adjuvants. In some embodiments, apharmaceutical composition described herein includes a second activeagent such as an additional therapeutic agent, (e.g., achemotherapeutic).

Formulations

Pharmaceutical compositions can be specially formulated foradministration in solid or liquid form, including those adapted for thefollowing: oral administration, for example, drenches (aqueous ornon-aqueous solutions or suspensions), tablets (e.g., those targeted forbuccal, sublingual, and systemic absorption), capsules, boluses,powders, granules, pastes for application to the tongue, andintraduodenal routes; parenteral administration, including intravenous,intraarterial, subcutaneous, intramuscular, intravascular,intraperitoneal or infusion as, for example, a sterile solution orsuspension, or sustained-release formulation; topical application, forexample, as a cream, ointment, or a controlled-release patch or sprayapplied to the skin; intravaginally or intrarectally, for example, as apessary, cream, stent or foam; sublingually; ocularly; pulmonarily;local delivery by catheter or stent; intrathecally, or nasally.

Examples of suitable aqueous and nonaqueous carriers which can beemployed in pharmaceutical compositions include water, ethanol, polyols(such as glycerol, propylene glycol, polyethylene glycol, and the like),and suitable mixtures thereof, vegetable oils, such as olive oil, andinjectable organic esters, such as ethyl oleate. Proper fluidity can bemaintained, for example, by the use of coating materials, such aslecithin, by the maintenance of the required particle size in the caseof dispersions, and by the use of surfactants.

These compositions can also contain adjuvants such as preservatives,wetting agents, emulsifying agents, dispersing agents, lubricants,and/or antioxidants. Prevention of the action of microorganisms upon thecompounds described herein can be ensured by the inclusion of variousantibacterial and antifungal agents, for example, paraben,chlorobutanol, phenol sorbic acid, and the like. It can also bedesirable to include isotonic agents, such as sugars, sodium chloride,and the like into the compositions. In addition, prolonged absorption ofthe injectable pharmaceutical form can be brought about by the inclusionof agents which delay absorption such as aluminum monostearate andgelatin.

Methods of preparing these formulations or compositions include the stepof bringing into association a compound described herein and/or thechemotherapeutic with the carrier and, optionally, one or more accessoryingredients. In general, the formulations are prepared by uniformly andintimately bringing into association a compound as disclosed herein withliquid carriers, or finely divided solid carriers, or both, and then, ifnecessary, shaping the product.

Preparations for such pharmaceutical compositions are well-known in theart. See, e.g., Anderson, Philip O.; Knoben, James E.; Troutman, WilliamG, eds., Handbook of Clinical Drug Data, Tenth Edition, McGraw-Hill,2002; Pratt and Taylor, eds., Principles of Drug Action, Third Edition,Churchill Livingston, N.Y., 1990; Katzung, ed., Basic and ClinicalPharmacology, Twelfth Edition, McGraw Hill, 2011; Goodman and Gilman,eds., The Pharmacological Basis of Therapeutics, Tenth Edition, McGrawHill, 2001; Remingtons Pharmaceutical Sciences, 20th Ed., LippincottWilliams & Wilkins., 2000; Martindale, The Extra Pharmacopoeia,Thirty-Second Edition (The Pharmaceutical Press, London, 1999); all ofwhich are incorporated by reference herein in their entirety. Exceptinsofar as any conventional excipient medium is incompatible with thecompounds provided herein, such as by producing any undesirablebiological effect or otherwise interacting in a deleterious manner withany other component(s) of the pharmaceutically acceptable composition,the excipient's use is contemplated to be within the scope of thisdisclosure.

In some embodiments, pharmaceutical compositions provided herein includeCD44 (cluster of differentiation 44) targeted nanoparticle formulations.CD44 is a transmembrane glycoprotein that plays a role in the cellsignaling cascades associated with cancer initiation and progression.CD44 is over-expressed on the surface of certain cancers, and hyaluronicacid has an affinity for these over-expressed CD44. Targeted drugdelivery system based on hyaluronic acid (e.g., hyaluronic acid modifiedmesoporous silica nanoparticles) can be an effective means to deliverdrugs to cancer cells. See, e.g., S. Arpicco, et al., Molecules, 2014,19, 3193-3230.

In some embodiments, the concentration of one or more of the compoundsprovided in the disclosed pharmaceutical compositions is less than about100%, about 90%, about 80%, about 70%, about 60%, about 50%, about 40%,about 30%, about 20%, about 19%, about 18%, about 17%, about 16%, about15%, about 14%, about 13%, about 12%, about 11%, about 10%, about 9%,about 8%, about 7%, about 6%, about 5%, about 4%, about 3%, about 2%,about 1%, about 0.5%, about 0.4%, about 0.3%, about 0.2%, about 0.1%,about 0.09%, about 0.08%, about 0.07%, about 0.06%, about 0.05%, about0.04%, about 0.03%, about 0.02%, about 0.01%, about 0.009%, about0.008%, about 0.007%, about 0.006%, about 0.005%, about 0.004%, about0.003%, about 0.002%, about 0.001%, about 0.0009%, about 0.0008%, about0.0007%, about 0.0006%, about 0.0005%, about 0.0004%, about 0.0003%,about 0.0002%, or about 0.0001%, w/w, w/v or v/v.

In some embodiments, the concentration of one or more of the compoundsas disclosed herein is greater than about 90%, about 80%, about 70%,about 60%, about 50%, about 40%, about 30%, about 20%, about 19.75%,about 19.50%, about 19.25%, about 19%, about 18.75%, about 18.50%, about18.25%, about 18%, about 17.75%, about 17.50%, about 17.25%, about 17%,about 16.75%, about 16.50%, about 16.25%, about 16%, about 15.75%, about15.50%, about 15.25%, about 15%, about 14.75%, about 14.50%, about14.25%, about 14%, about 13.75%, about 13.50%, about 13.25%, about 13%,about 12.75%, about 12.50%, about 12.25%, about 12%, about 11.75%, about11.50%, about 11.25%, about 11%, about 10.75%, about 10.50%, about10.25%, about 10%, about 9.75%, about 9.50%, about 9.25%, about 9%,about 8.75%, about 8.50%, about 8.25%, about 8%, about 7.75%, about7.50%, about 7.25%, about 7%, about 6.75%, about 6.50%, about 6.25%,about 6%, about 5.75%, about 5.50%, about 5.25%, about 5%, about 4.75%,about 4.50%, about 4.25%, about 4%, about 3.75%, about 3.50%, about3.25%, about 3%, about 2.75%, about 2.50%, about 2.25%, about 2%, about1.75%, about 1.50%, about 1.25%, about 1%, about 0.5%, about 0.4%, about0.3%, about 0.2%, about 0.1%, about 0.09%, about 0.08%, about 0.07%,about 0.06%, about 0.05%, about 0.04%, about 0.03%, about 0.02%, about0.01%, about 0.009%, about 0.008%, about 0.007%, about 0.006%, about0.005%, about 0.004%, about 0.003%, about 0.002%, about 0.001%, about0.0009%, about 0.0008%, about 0.0007%, about 0.0006%, about 0.0005%,about 0.0004%, about 0.0003%, about 0.0002%, or about 0.0001%, w/w, w/v,or v/v.

In some embodiments, the concentration of one or more of the compoundsas disclosed herein is in the range from approximately 0.0001% toapproximately 50%, approximately 0.001% to approximately 40%,approximately 0.01% to approximately 30%, approximately 0.02% toapproximately 29%, approximately 0.03% to approximately 28%,approximately 0.04% to approximately 27%, approximately 0.05% toapproximately 26%, approximately 0.06% to approximately 25%,approximately 0.07% to approximately 24%, approximately 0.08% toapproximately 23%, approximately 0.09% to approximately 22%,approximately 0.1% to approximately 21%, approximately 0.2% toapproximately 20%, approximately 0.3% to approximately 19%,approximately 0.4% to approximately 18%, approximately 0.5% toapproximately 17%, approximately 0.6% to approximately 16%,approximately 0.7% to approximately 15%, approximately 0.8% toapproximately 14%, approximately 0.9% to approximately 12%, orapproximately 1% to approximately 10%, w/w, w/v or v/v.

In some embodiments, the concentration of one or more of the compoundsas disclosed herein is in the range from approximately 0.001% toapproximately 10%, approximately 0.01% to approximately 5%,approximately 0.02% to approximately 4.5%, approximately 0.03% toapproximately 4%, approximately 0.04% to approximately 3.5%,approximately 0.05% to approximately 3%, approximately 0.06% toapproximately 2.5%, approximately 0.07% to approximately 2%,approximately 0.08% to approximately 1.5%, approximately 0.09% toapproximately 1%, or approximately 0.1% to approximately 0.9%, w/w, w/vor v/v.

In some embodiments, the amount of one or more of the compounds asdisclosed herein is equal to or less than about 10 g, about 9.5 g, about9.0 g, about 8.5 g, about 8.0 g, about 7.5 g, about 7.0 g, about 6.5 g,about 6.0 g, about 5.5 g, about 5.0 g, about 4.5 g, about 4.0 g, about3.5 g, about 3.0 g, about 2.5 g, about 2.0 g, about 1.5 g, about 1.0 g,about 0.95 g, about 0.9 g, about 0.85 g, about 0.8 g, about 0.75 g,about 0.7 g, about 0.65 g, about 0.6 g, about 0.55 g, about 0.5 g, about0.45 g, about 0.4 g, about 0.35 g, about 0.3 g, about 0.25 g, about 0.2g, about 0.15 g, about 0.1 g, about 0.09 g, about 0.08 g, about 0.07 g,about 0.06 g, about 0.05 g, about 0.04 g, about 0.03 g, about 0.02 g,about 0.01 g, about 0.009 g, about 0.008 g, about 0.007 g, about 0.006g, about 0.005 g, about 0.004 g, about 0.003 g, about 0.002 g, about0.001 g, about 0.0009 g, about 0.0008 g, about 0.0007 g, about 0.0006 g,about 0.0005 g, about 0.0004 g, about 0.0003 g, about 0.0002 g, or about0.0001 g.

In some embodiments, the amount of one or more of the compounds asdisclosed herein is more than about 0.0001 g, about 0.0002 g, about0.0003 g, about 0.0004 g, about 0.0005 g, about 0.0006 g, about 0.0007g, about 0.0008 g, about 0.0009 g, about 0.001 g, about 0.0015 g, about0.002 g, about 0.0025 g, about 0.003 g, about 0.0035 g, about 0.004 g,about 0.0045 g, about 0.005 g, about 0.0055 g, about 0.006 g, about0.0065 g, about 0.007 g, about 0.0075 g, about 0.008 g, about 0.0085 g,about 0.009 g, about 0.0095 g, about 0.01 g, about 0.015 g, about 0.02g, about 0.025 g, about 0.03 g, about 0.035 g, about 0.04 g, about 0.045g, about 0.05 g, about 0.055 g, about 0.06 g, about 0.065 g, about 0.07g, about 0.075 g, about 0.08 g, about 0.085 g, about 0.09 g, about 0.095g, about 0.1 g, about 0.15 g, about 0.2 g, about 0.25 g, about 0.3 g,about 0.35 g, about 0.4 g, about 0.45 g, about 0.5 g, about 0.55 g,about 0.6 g, about 0.65 g, about 0.7 g, about 0.75 g, about 0.8 g, about0.85 g, about 0.9 g, about 0.95 g, about 1 g, about 1.5 g, about 2 g,about 2.5 g, about 3 g, about 3.5 g, about 4 g, about 4.5 g, about 5 g,about 5.5 g, about 6 g, about 6.5 g, about 7 g, about 7.5 g, about 8 g,about 8.5 g, about 9 g, about 9.5 g, or about 10 g.

In some embodiments, the amount of one or more of the compounds asdisclosed herein is in the range of about 0.0001 to about 10 g, about0.0005 to about 9 g, about 0.001 to about 8 g, about 0.005 to about 7 g,about 0.01 to about 6 g, about 0.05 to about 5 g, about 0.1 to about 4g, about 0.5 to about 4 g, or about 1 to about 3 g.

1A. Formulations for Oral Administration

In some embodiments, provided herein are pharmaceutical compositions fororal administration containing a compound as disclosed herein, and apharmaceutical excipient suitable for oral administration. In someembodiments, provided herein are pharmaceutical compositions for oraladministration containing: (i) an effective amount of a disclosedcompound; optionally (ii) an effective amount of one or more secondagents; and (iii) one or more pharmaceutical excipients suitable fororal administration. In some embodiments, the pharmaceutical compositionfurther contains: (iv) an effective amount of a third agent.

In some embodiments, the pharmaceutical composition can be a liquidpharmaceutical composition suitable for oral consumption. Pharmaceuticalcompositions suitable for oral administration can be presented asdiscrete dosage forms, such as capsules, cachets, or tablets, or liquidsor aerosol sprays each containing a predetermined amount of an activeingredient as a powder or in granules, a solution, or a suspension in anaqueous or non-aqueous liquid, an oil-in-water emulsion, or awater-in-oil liquid emulsion. Such dosage forms can be prepared by anyof the methods of pharmacy, but all methods include the step of bringingthe active ingredient into association with the carrier, whichconstitutes one or more ingredients. In general, the pharmaceuticalcompositions are prepared by uniformly and intimately admixing theactive ingredient with liquid carriers or finely divided solid carriersor both, and then, if necessary, shaping the product into the desiredpresentation. For example, a tablet can be prepared by compression ormolding, optionally with one or more accessory ingredients. Compressedtablets can be prepared by compressing in a suitable machine the activeingredient in a free-flowing form such as powder or granules, optionallymixed with an excipient such as, but not limited to, a binder, alubricant, an inert diluent, and/or a surface active or dispersingagent. Molded tablets can be made by molding in a suitable machine amixture of the powdered compound moistened with an inert liquid diluent.

The present disclosure further encompasses anhydrous pharmaceuticalcompositions and dosage forms comprising an active ingredient, sincewater can facilitate the degradation of some compounds. For example,water can be added (e.g., about 5%) in the pharmaceutical arts as ameans of simulating long-term storage in order to determinecharacteristics such as shelf-life or the stability of formulations overtime. Anhydrous pharmaceutical compositions and dosage forms can beprepared using anhydrous or low moisture containing ingredients and lowmoisture or low humidity conditions. For example, pharmaceuticalcompositions and dosage forms which contain lactose can be madeanhydrous if substantial contact with moisture and/or humidity duringmanufacturing, packaging, and/or storage is expected. An anhydrouspharmaceutical composition can be prepared and stored such that itsanhydrous nature is maintained. Accordingly, anhydrous pharmaceuticalcompositions can be packaged using materials known to prevent exposureto water such that they can be included in suitable formulary kits.Examples of suitable packaging include, but are not limited to,hermetically sealed foils, plastic or the like, unit dose containers,blister packs, and strip packs.

An active ingredient can be combined in an intimate admixture with apharmaceutical carrier according to conventional pharmaceuticalcompounding techniques. The carrier can take a wide variety of formsdepending on the form of preparation desired for administration. Inpreparing the pharmaceutical compositions for an oral dosage form, anyof the usual pharmaceutical media can be employed as carriers, such as,for example, water, glycols, oils, alcohols, flavoring agents,preservatives, coloring agents, and the like in the case of oral liquidpreparations (such as suspensions, solutions, and elixirs) or aerosols;or carriers such as starches, sugars, micro-crystalline cellulose,diluents, granulating agents, lubricants, binders, and disintegratingagents can be used in the case of oral solid preparations, in someembodiments without employing the use of lactose. For example, suitablecarriers include powders, capsules, and tablets, with the solid oralpreparations. In some embodiments, tablets can be coated by standardaqueous or nonaqueous techniques.

Binders suitable for use in pharmaceutical compositions and dosage formsinclude, but are not limited to, corn starch, potato starch, or otherstarches, gelatin, natural and synthetic gums such as acacia, sodiumalginate, alginic acid, other alginates, powdered tragacanth, guar gum,cellulose and its derivatives (e.g., ethyl cellulose, cellulose acetate,carboxymethyl cellulose calcium, sodium carboxymethyl cellulose),polyvinyl pyrrolidone, methyl cellulose, pre-gelatinized starch,hydroxypropyl methyl cellulose, microcrystalline cellulose, and mixturesthereof.

Examples of suitable fillers for use in the pharmaceutical compositionsand dosage forms disclosed herein include, but are not limited to, talc,calcium carbonate (e.g., granules or powder), microcrystallinecellulose, powdered cellulose, dextrates, kaolin, mannitol, silicicacid, sorbitol, starch, pre-gelatinized starch, and mixtures thereof.

Disintegrants can be used in the pharmaceutical compositions as providedherein to provide tablets that disintegrate when exposed to an aqueousenvironment. Too much of a disintegrant can produce tablets which candisintegrate in the bottle. Too little can be insufficient fordisintegration to occur and can thus alter the rate and extent ofrelease of the active ingredient(s) from the dosage form. Thus, asufficient amount of disintegrant that is neither too little nor toomuch to detrimentally alter the release of the active ingredient(s) canbe used to form the dosage forms of the compounds disclosed herein. Theamount of disintegrant used can vary based upon the type of formulationand mode of administration, and can be readily discernible to those ofordinary skill in the art. About 0.5 to about 15 weight percent ofdisintegrant, or about 1 to about 5 weight percent of disintegrant, canbe used in the pharmaceutical composition. Disintegrants that can beused to form pharmaceutical compositions and dosage forms include, butare not limited to, agar-agar, alginic acid, calcium carbonate,microcrystalline cellulose, croscarmellose sodium, crospovidone,polacrilin potassium, sodium starch glycolate, potato or tapioca starch,other starches, pre-gelatinized starch, other starches, clays, otheralgins, other celluloses, gums or mixtures thereof.

Lubricants which can be used to form pharmaceutical compositions anddosage forms include, but are not limited to, calcium stearate,magnesium stearate, mineral oil, light mineral oil, glycerin, sorbitol,mannitol, polyethylene glycol, other glycols, stearic acid, sodiumlauryl sulfate, talc, hydrogenated vegetable oil (e.g., peanut oil,cottonseed oil, sunflower oil, sesame oil, olive oil, corn oil, andsoybean oil), zinc stearate, ethyl oleate, ethylaureate, agar, ormixtures thereof. Additional lubricants include, for example, a syloidsilica gel, a coagulated aerosol of synthetic silica, or mixturesthereof. A lubricant can optionally be added, in an amount of less thanabout 1 weight percent of the pharmaceutical composition.

When aqueous suspensions and/or elixirs are desired for oraladministration, the active ingredient therein can be combined withvarious sweetening or flavoring agents, coloring matter or dyes and, forexample, emulsifying and/or suspending agents, together with suchdiluents as water, ethanol, propylene glycol, glycerin and variouscombinations thereof.

The tablets can be uncoated or coated by known techniques to delaydisintegration and absorption in the gastrointestinal tract and therebyprovide a sustained action over a longer period. For example, a timedelay material such as glyceryl monostearate or glyceryl distearate canbe employed. Formulations for oral use can also be presented as hardgelatin capsules wherein the active ingredient is mixed with an inertsolid diluent, for example, calcium carbonate, calcium phosphate orkaolin, or as soft gelatin capsules wherein the active ingredient ismixed with water or an oil medium, for example, peanut oil, liquidparaffin or olive oil.

Surfactant which can be used to form pharmaceutical compositions anddosage forms include, but are not limited to, hydrophilic surfactants,lipophilic surfactants, and mixtures thereof. That is, a mixture ofhydrophilic surfactants can be employed, a mixture of lipophilicsurfactants can be employed, or a mixture of at least one hydrophilicsurfactant and at least one lipophilic surfactant can be employed.

A suitable hydrophilic surfactant can generally have an HLB value of atleast about 10, while suitable lipophilic surfactants can generally havean HLB value of or less than about 10. An empirical parameter used tocharacterize the relative hydrophilicity and hydrophobicity of non-ionicamphiphilic compounds is the hydrophilic-lipophilic balance (“HLB”value). Surfactants with lower HLB values are more lipophilic orhydrophobic, and have greater solubility in oils, while surfactants withhigher HLB values are more hydrophilic, and have greater solubility inaqueous solutions. Hydrophilic surfactants are generally considered tobe those compounds having an HLB value greater than about 10, as well asanionic, cationic, or zwitterionic compounds for which the HLB scale isnot generally applicable. Similarly, lipophilic (i.e., hydrophobic)surfactants are compounds having an HLB value equal to or less thanabout 10. However, HLB value of a surfactant is merely a rough guidegenerally used to enable formulation of industrial, pharmaceutical andcosmetic emulsions.

Hydrophilic surfactants can be either ionic or non-ionic. Suitable ionicsurfactants include, but are not limited to, alkylammonium salts;fusidic acid salts; fatty acid derivatives of amino acids,oligopeptides, and polypeptides; glyceride derivatives of amino acids,oligopeptides, and polypeptides; lecithins and hydrogenated lecithins;lysolecithins and hydrogenated lysolecithins; phospholipids andderivatives thereof; lysophospholipids and derivatives thereof;carnitine fatty acid ester salts; salts of alkylsulfates; fatty acidsalts; sodium docusate; acylactylates; mono- and di-acetylated tartaricacid esters of mono- and di-glycerides; succinylated mono- anddi-glycerides; citric acid esters of mono- and di-glycerides; andmixtures thereof.

Within the aforementioned group, ionic surfactants include, by way ofexample: lecithins, lysolecithin, phospholipids, lysophospholipids andderivatives thereof; carnitine fatty acid ester salts; salts ofalkylsulfates; fatty acid salts; sodium docusate; acylactylates; mono-and di-acetylated tartaric acid esters of mono- and di-glycerides;succinylated mono- and di-glycerides; citric acid esters of mono- anddi-glycerides; and mixtures thereof.

Ionic surfactants can be the ionized forms of lecithin, lysolecithin,phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol,phosphatidic acid, phosphatidylserine, lysophosphatidylcholine,lysophosphatidylethanolamine, lysophosphatidylglycerol, lysophosphatidicacid, lysophosphatidylserine, PEG-phosphatidylethanolamine,PVP-phosphatidylethanolamine, lactylic esters of fatty acids,stearoyl-2-lactylate, stearoyl lactylate, succinylated monoglycerides,mono/diacetylated tartaric acid esters of mono/diglycerides, citric acidesters of mono/diglycerides, cholylsarcosine, caproate, caprylate,caprate, laurate, myristate, palmitate, oleate, ricinoleate, linoleate,linolenate, stearate, lauryl sulfate, teracecyl sulfate, docusate,lauroyl carnitines, palmitoyl carnitines, myristoyl carnitines, andsalts and mixtures thereof.

Hydrophilic non-ionic surfactants can include, but are not limited to,alkylglucosides; alkylmaltosides; alkylthioglucosides; laurylmacrogolglycerides; polyoxyalkylene alkyl ethers such as polyethyleneglycol alkyl ethers; polyoxyalkylene alkylphenols such as polyethyleneglycol alkyl phenols; polyoxyalkylene alkyl phenol fatty acid esterssuch as polyethylene glycol fatty acids monoesters and polyethyleneglycol fatty acids diesters; polyethylene glycol glycerol fatty acidesters; polyglycerol fatty acid esters; polyoxyalkylene sorbitan fattyacid esters such as polyethylene glycol sorbitan fatty acid esters;hydrophilic transesterification products of a polyol with at least onemember of glycerides, vegetable oils, hydrogenated vegetable oils, fattyacids, and sterols; polyoxyethylene sterols, derivatives, and analoguesthereof; polyoxyethylated vitamins and derivatives thereof;polyoxyethylene-polyoxypropylene block copolymers; and mixtures thereof;polyethylene glycol sorbitan fatty acid esters and hydrophilictransesterification products of a polyol with at least one member oftriglycerides, vegetable oils, and hydrogenated vegetable oils. Thepolyol can be glycerol, ethylene glycol, polyethylene glycol, sorbitol,propylene glycol, pentaerythritol, or a saccharide.

Other hydrophilic-non-ionic surfactants include, without limitation,PEG-10 laurate, PEG-12 laurate, PEG-20 laurate, PEG-32 laurate, PEG-32dilaurate, PEG-12 oleate, PEG-15 oleate, PEG-20 oleate, PEG-20 dioleate,PEG-32 oleate, PEG-200 oleate, PEG-400 oleate, PEG-15 stearate, PEG-32distearate, PEG-40 stearate, PEG-100 stearate, PEG-20 dilaurate, PEG-25glyceryl trioleate, PEG-32 dioleate, PEG-20 glyceryl laurate, PEG-30glyceryl laurate, PEG-20 glyceryl stearate, PEG-20 glyceryl oleate,PEG-30 glyceryl oleate, PEG-30 glyceryl laurate, PEG-40 glyceryllaurate, PEG-40 palm kernel oil, PEG-50 hydrogenated castor oil, PEG-40castor oil, PEG-35 castor oil, PEG-60 castor oil, PEG-40 hydrogenatedcastor oil, PEG-60 hydrogenated castor oil, PEG-60 corn oil, PEG-6caprate/caprylate glycerides, PEG-8 caprate/caprylate glycerides,polyglyceryl-10 laurate, PEG-30 cholesterol, PEG-25 phyto sterol, PEG-30soya sterol, PEG-20 trioleate, PEG-40 sorbitan oleate, PEG-80 sorbitanlaurate, polysorbate 20, polysorbate 80, POE-9 lauryl ether, POE-23lauryl ether, POE-10 oleyl ether, POE-20 oleyl ether, POE-20 stearylether, tocopheryl PEG-100 succinate, PEG-24 cholesterol, polyglyceryl-10oleate, Tween 40, Tween 60, sucrose monostearate, sucrose monolaurate,sucrose monopalmitate, PEG 10-100 nonyl phenol series, PEG 15-100 octylphenol series, and poloxamers.

Suitable lipophilic surfactants include, by way of example only: fattyalcohols; glycerol fatty acid esters; acetylated glycerol fatty acidesters; lower alcohol fatty acids esters; propylene glycol fatty acidesters; sorbitan fatty acid esters; polyethylene glycol sorbitan fattyacid esters; sterols and sterol derivatives; polyoxyethylated sterolsand sterol derivatives; polyethylene glycol alkyl ethers; sugar esters;sugar ethers; lactic acid derivatives of mono- and di-glycerides;hydrophobic transesterification products of a polyol with at least onemember of glycerides, vegetable oils, hydrogenated vegetable oils, fattyacids and sterols; oil-soluble vitamins/vitamin derivatives; andmixtures thereof. Within this group, non-limiting examples of lipophilicsurfactants include glycerol fatty acid esters, propylene glycol fattyacid esters, and mixtures thereof, or are hydrophobictransesterification products of a polyol with at least one member ofvegetable oils, hydrogenated vegetable oils, and triglycerides.

In one embodiment, the pharmaceutical composition can include asolubilizer to ensure good solubilization and/or dissolution of acompound as provided herein and to minimize precipitation of thecompound. This can be especially important for pharmaceuticalcompositions for non-oral use, e.g., pharmaceutical compositions forinjection. A solubilizer can also be added to increase the solubility ofthe hydrophilic drug and/or other components, such as surfactants, or tomaintain the pharmaceutical composition as a stable or homogeneoussolution or dispersion.

Examples of suitable solubilizers include, but are not limited to, thefollowing: alcohols and polyols, such as ethanol, isopropanol, butanol,benzyl alcohol, ethylene glycol, propylene glycol, butanediols andisomers thereof, glycerol, pentaerythritol, sorbitol, mannitol,transcutol, dimethyl isosorbide, polyethylene glycol, polypropyleneglycol, polyvinylalcohol, hydroxypropyl methylcellulose and othercellulose derivatives, cyclodextrins and cyclodextrin derivatives;ethers of polyethylene glycols having an average molecular weight ofabout 200 to about 6000, such as tetrahydrofurfuryl alcohol PEG ether(glycofurol) or methoxy PEG; amides and other nitrogen-containingcompounds such as 2-pyrrolidone, 2-piperidone, ε-caprolactam,N-alkylpyrrolidone, N-hydroxyalkylpyrrolidone, N-alkylpiperidone,N-alkylcaprolactam, dimethylacetamide and polyvinylpyrrolidone; esterssuch as ethyl propionate, tributylcitrate, acetyl triethylcitrate,acetyl tributyl citrate, triethylcitrate, ethyl oleate, ethyl caprylate,ethyl butyrate, triacetin, propylene glycol monoacetate, propyleneglycol diacetate, ε-caprolactone and isomers thereof, 6-valerolactoneand isomers thereof, β-butyrolactone and isomers thereof; and othersolubilizers known in the art, such as dimethyl acetamide, dimethylisosorbide, N-methyl pyrrolidones, monooctanoin, diethylene glycolmonoethyl ether, and water.

Mixtures of solubilizers can also be used. Examples include, but notlimited to, triacetin, triethylcitrate, ethyl oleate, ethyl caprylate,dimethylacetamide, N-methylpyrrolidone, N-hydroxyethylpyrrolidone,polyvinylpyrrolidone, hydroxypropyl methylcellulose, hydroxypropylcyclodextrins, ethanol, polyethylene glycol 200-100, glycofurol,transcutol, propylene glycol, and dimethyl isosorbide. In someembodiments, solubilizers include sorbitol, glycerol, triacetin, ethylalcohol, PEG-400, glycofurol and propylene glycol.

The amount of solubilizer that can be included is not particularlylimited. The amount of a given solubilizer can be limited to abioacceptable amount, which can be readily determined by one of skill inthe art. In some circumstances, it can be advantageous to includeamounts of solubilizers far in excess of bioacceptable amounts, forexample to maximize the concentration of the drug, with excesssolubilizer removed prior to providing the pharmaceutical composition toa subject using conventional techniques, such as distillation orevaporation. Thus, if present, the solubilizer can be in a weight ratioof about 10%, 25%, 50%, 100%, or up to about 200% by weight, based onthe combined weight of the drug, and other excipients. If desired, verysmall amounts of solubilizer can also be used, such as about 5%, 2%, 1%or even less. Typically, the solubilizer can be present in an amount ofabout 1% to about 100%, more typically about 5% to about 25% by weight.

The pharmaceutical composition can further include one or morepharmaceutically acceptable additives and excipients. Such additives andexcipients include, without limitation, detackifiers, anti-foamingagents, buffering agents, polymers, antioxidants, preservatives,chelating agents, viscomodulators, tonicifiers, flavorants, colorants,oils, odorants, opacifiers, suspending agents, binders, fillers,plasticizers, lubricants, and mixtures thereof.

Exemplary preservatives can include antioxidants, chelating agents,antimicrobial preservatives, antifungal preservatives, alcoholpreservatives, acidic preservatives, and other preservatives. Exemplaryantioxidants include, but are not limited to, alpha tocopherol, ascorbicacid, acorbyl palmitate, butylated hydroxyanisole, butylatedhydroxytoluene, monothioglycerol, potassium metabisulfite, propionicacid, propyl gallate, sodium ascorbate, sodium bisulfite, sodiummetabisulfite, and sodium sulfite. Exemplary chelating agents includeethylenediaminetetraacetic acid (EDTA), citric acid monohydrate,disodium edetate, dipotassium edetate, edetic acid, fumaric acid, malicacid, phosphoric acid, sodium edetate, tartaric acid, and trisodiumedetate. Exemplary antimicrobial preservatives include, but are notlimited to, benzalkonium chloride, benzethonium chloride, benzylalcohol, bronopol, cetrimide, cetylpyridinium chloride, chlorhexidine,chlorobutanol, chlorocresol, chloroxylenol, cresol, ethyl alcohol,glycerin, hexetidine, imidurea, phenol, phenoxyethanol, phenylethylalcohol, phenylmercuric nitrate, propylene glycol, and thimerosal.Exemplary antifungal preservatives include, but are not limited to,butyl paraben, methyl paraben, ethyl paraben, propyl paraben, benzoicacid, hydroxybenzoic acid, potassium benzoate, potassium sorbate, sodiumbenzoate, sodium propionate, and sorbic acid. Exemplary alcoholpreservatives include, but are not limited to, ethanol, polyethyleneglycol, phenol, phenolic compounds, bisphenol, chlorobutanol,hydroxybenzoate, and phenylethyl alcohol. Exemplary acidic preservativesinclude, but are not limited to, vitamin A, vitamin C, vitamin E,beta-carotene, citric acid, acetic acid, dehydroacetic acid, ascorbicacid, sorbic acid, and phytic acid. Other preservatives include, but arenot limited to, tocopherol, tocopherol acetate, deteroxime mesylate,cetrimide, butylated hydroxyanisol (BHA), butylated hydroxytoluened(BHT), ethylenediamine, sodium lauryl sulfate (SLS), sodium lauryl ethersulfate (SLES), sodium bisulfite, sodium metabisulfite, potassiumsulfite, potassium metabisulfite, Glydant Plus, Phenonip, methylparaben,Germall 115, Germaben II, Neolone, Kathon, and Euxyl. In certainembodiments, the preservative is an anti-oxidant. In other embodiments,the preservative is a chelating agent.

Exemplary oils include, but are not limited to, almond, apricot kernel,avocado, babassu, bergamot, black current seed, borage, cade, camomile,canola, caraway, carnauba, castor, cinnamon, cocoa butter, coconut, codliver, coffee, corn, cotton seed, emu, eucalyptus, evening primrose,fish, flaxseed, geraniol, gourd, grape seed, hazel nut, hyssop,isopropyl myristate, jojoba, kukui nut, lavandin, lavender, lemon,litsea cubeba, macademia nut, mallow, mango seed, meadowfoam seed, mink,nutmeg, olive, orange, orange roughy, palm, palm kernel, peach kernel,peanut, poppy seed, pumpkin seed, rapeseed, rice bran, rosemary,safflower, sandalwood, sasquana, savoury, sea buckthorn, sesame, sheabutter, silicone, soybean, sunflower, tea tree, thistle, tsubaki,vetiver, walnut, and wheat germ oils. Exemplary oils include, but arenot limited to, butyl stearate, caprylic triglyceride, caprictriglyceride, cyclomethicone, diethyl sebacate, dimethicone 360,isopropyl myristate, mineral oil, octyldodecanol, oleyl alcohol,silicone oil, and combinations thereof.

In addition, an acid or a base can be incorporated into thepharmaceutical composition to facilitate processing, to enhancestability, or for other reasons. Examples of pharmaceutically acceptablebases include amino acids, amino acid esters, ammonium hydroxide,potassium hydroxide, sodium hydroxide, sodium hydrogen carbonate,aluminum hydroxide, calcium carbonate, magnesium hydroxide, magnesiumaluminum silicate, synthetic aluminum silicate, synthetic hydrocalcite,magnesium aluminum hydroxide, diisopropylethylamine, ethanolamine,ethylenediamine, triethanolamine, triethylamine, triisopropanolamine,trimethylamine, tris(hydroxymethyl)aminomethane (TRIS) and the like.Also suitable are bases that are salts of a pharmaceutically acceptableacid, such as acetic acid, acrylic acid, adipic acid, alginic acid,alkanesulfonic acid, amino acids, ascorbic acid, benzoic acid, boricacid, butyric acid, carbonic acid, citric acid, fatty acids, formicacid, fumaric acid, gluconic acid, hydroquinosulfonic acid, isoascorbicacid, lactic acid, maleic acid, oxalic acid, para-bromophenylsulfonicacid, propionic acid, p-toluenesulfonic acid, salicylic acid, stearicacid, succinic acid, tannic acid, tartaric acid, thioglycolic acid,toluenesulfonic acid, uric acid, and the like. Salts of polyproticacids, such as sodium phosphate, disodium hydrogen phosphate, and sodiumdihydrogen phosphate can also be used. When the base is a salt, thecation can be any convenient and pharmaceutically acceptable cation,such as ammonium, alkali metals, alkaline earth metals, and the like.Examples can include, but not limited to, sodium, potassium, lithium,magnesium, calcium and ammonium.

Suitable acids are pharmaceutically acceptable organic or inorganicacids. Examples of suitable inorganic acids include hydrochloric acid,hydrobromic acid, hydriodic acid, sulfuric acid, nitric acid, boricacid, phosphoric acid, and the like. Examples of suitable organic acidsinclude acetic acid, acrylic acid, adipic acid, alginic acid,alkanesulfonic acids, amino acids, ascorbic acid, benzoic acid, boricacid, butyric acid, carbonic acid, citric acid, fatty acids, formicacid, fumaric acid, gluconic acid, hydroquinosulfonic acid, isoascorbicacid, lactic acid, maleic acid, methanesulfonic acid, oxalic acid,para-bromophenylsulfonic acid, propionic acid, p-toluenesulfonic acid,salicylic acid, stearic acid, succinic acid, tannic acid, tartaric acid,thioglycolic acid, toluenesulfonic acid, uric acid and the like.

1B. Formulations for Parenteral Administration

In some embodiments, provided herein are pharmaceutical compositions forparenteral administration containing a compound as disclosed herein, anda pharmaceutical excipient suitable for parenteral administration. Insome embodiments, provided herein are pharmaceutical compositions forparenteral administration containing: (i) an effective amount of adisclosed compound; optionally (ii) an effective amount of one or moresecond agents; and (iii) one or more pharmaceutical excipients suitablefor parenteral administration. In some embodiments, the pharmaceuticalcomposition further contains: (iv) an effective amount of a third agent.

The forms in which the disclosed pharmaceutical compositions can beincorporated for administration by injection include aqueous or oilsuspensions, or emulsions, with sesame oil, corn oil, cottonseed oil, orpeanut oil, as well as elixirs, mannitol, dextrose, or a sterile aqueoussolution, and similar pharmaceutical vehicles.

Aqueous solutions in saline are also conventionally used for injection.Ethanol, glycerol, propylene glycol, liquid polyethylene glycol, and thelike (and suitable mixtures thereof), cyclodextrin derivatives, andvegetable oils can also be employed.

Aqueous solutions in saline are also conventionally used for injection.Ethanol, glycerol, propylene glycol, liquid polyethylene glycol, and thelike (and suitable mixtures thereof), cyclodextrin derivatives, andvegetable oils can also be employed. The proper fluidity can bemaintained, for example, by the use of a coating, such as lecithin, forthe maintenance of the required particle size in the case of dispersionand by the use of surfactants. The prevention of the action ofmicroorganisms can be brought about by various antibacterial andantifungal agents, for example, parabens, chlorobutanol, phenol, sorbicacid, thimerosal, and the like.

Sterile injectable solutions are prepared by incorporating a compound asdisclosed herein in the required amount in the appropriate solvent withvarious other ingredients as enumerated above, as appropriate, followedby filtered sterilization. Generally, dispersions are prepared byincorporating the various sterilized active ingredients into a sterilevehicle which contains the basic dispersion medium and the appropriateother ingredients from those enumerated above. In the case of sterilepowders for the preparation of sterile injectable solutions, certainmethods of preparation are vacuum-drying and freeze-drying techniqueswhich yield a powder of the active ingredient plus any additionalingredient from a previously sterile-filtered solution thereof.

The injectable formulations can be sterilized, for example, byfiltration through a bacterial-retaining filter, or by incorporatingsterilizing agents in the form of sterile solid compositions which canbe dissolved or dispersed in sterile water or other sterile injectablemedium prior to use. Injectable compositions can contain from about 0.1to about 5% w/w of a compound as disclosed herein.

1C. Formulations for Topical Administration

In some embodiments, provided herein are pharmaceutical compositions fortopical (e.g., transdermal) administration containing a compound asdisclosed herein, and a pharmaceutical excipient suitable for topicaladministration. In some embodiments, provided herein are pharmaceuticalcompositions for topical administration containing: (i) an effectiveamount of a disclosed compound; optionally (ii) an effective amount ofone or more second agents; and (iii) one or more pharmaceuticalexcipients suitable for topical administration. In some embodiments, thepharmaceutical composition further contains: (iv) an effective amount ofa third agent.

Pharmaceutical compositions provided herein can be formulated intopreparations in solid, semi-solid, or liquid forms suitable for local ortopical administration, such as gels, water soluble jellies, creams,lotions, suspensions, foams, powders, slurries, ointments, solutions,oils, pastes, suppositories, sprays, emulsions, saline solutions,dimethylsulfoxide (DMSO)-based solutions. In general, carriers withhigher densities are capable of providing an area with a prolongedexposure to the active ingredients. In contrast, a solution formulationcan provide more immediate exposure of the active ingredient to thechosen area.

The pharmaceutical compositions also can comprise suitable solid or gelphase carriers or excipients, which are compounds that allow increasedpenetration of, or assist in the delivery of, therapeutic moleculesacross the stratum corneum permeability barrier of the skin. There aremany of these penetration-enhancing molecules known to those trained inthe art of topical formulation. Examples of such carriers and excipientsinclude, but are not limited to, humectants (e.g., urea), glycols (e.g.,propylene glycol), alcohols (e.g., ethanol), fatty acids (e.g., oleicacid), surfactants (e.g., isopropyl myristate and sodium laurylsulfate), pyrrolidones, glycerol monolaurate, sulfoxides, terpenes(e.g., menthol), amines, amides, alkanes, alkanols, water, calciumcarbonate, calcium phosphate, various sugars, starches, cellulosederivatives, gelatin, and polymers such as polyethylene glycols.

Another exemplary formulation for use in the disclosed methods employstransdermal delivery devices (“patches”). Such transdermal patches canbe used to provide continuous or discontinuous infusion of a compound asprovided herein in controlled amounts, either with or without anotheragent.

The construction and use of transdermal patches for the delivery ofpharmaceutical agents is well known in the art. See, e.g., U.S. Pat.Nos. 5,023,252, 4,992,445 and 5,001,139. Such patches can be constructedfor continuous, pulsatile, or on demand delivery of pharmaceuticalagents.

Suitable devices for use in delivering intradermal pharmaceuticallyacceptable compositions described herein include short needle devicessuch as those described in U.S. Pat. Nos. 4,886,499; 5,190,521;5,328,483; 5,527,288; 4,270,537; 5,015,235; 5,141,496; and 5,417,662.Intradermal compositions can be administered by devices which limit theeffective penetration length of a needle into the skin, such as thosedescribed in PCT publication WO 99/34850 and functional equivalentsthereof. Jet injection devices which deliver liquid vaccines to thedermis via a liquid jet injector and/or via a needle which pierces thestratum corneum and produces a jet which reaches the dermis aresuitable. Jet injection devices are described, for example, in U.S. Pat.Nos. 5,480,381; 5,599,302; 5,334,144; 5,993,412; 5,649,912; 5,569,189;5,704,911; 5,383,851; 5,893,397; 5,466,220; 5,339,163; 5,312,335;5,503,627; 5,064,413; 5,520,639; 4,596,556; 4,790,824; 4,941,880;4,940,460; and PCT publications WO 97/37705 and WO 97/13537. Ballisticpowder/particle delivery devices which use compressed gas to acceleratevaccine in powder form through the outer layers of the skin to thedermis are suitable. Alternatively or additionally, conventionalsyringes can be used in the classical mantoux method of intradermaladministration.

Topically-administrable formulations can, for example, comprise fromabout 1% to about 10% (w/w) of a compound provided herein relative tothe total weight of the formulation, although the concentration of thecompound provided herein in the formulation can be as high as thesolubility limit of the compound in the solvent. In some embodiments,topically-administrable formulations can, for example, comprise fromabout 1% to about 9% (w/w) of a compound provided herein, such as fromabout 1% to about 8% (w/w), further such as from about 1% to about 7%(w/w), further such as from about 1% to about 6% (w/w), further such asfrom about 1% to about 5% (w/w), further such as from about 1% to about4% (w/w), further such as from about 1% to about 3% (w/w), and furthersuch as from about 1% to about 2% (w/w) of a compound provided herein.Formulations for topical administration can further comprise one or moreof the additional pharmaceutically acceptable excipients describedherein.

1D. Formulations for Inhalation Administration

In some embodiments, provided herein are pharmaceutical compositions forinhalation administration containing a compound as disclosed herein, anda pharmaceutical excipient suitable for topical administration. In someembodiments, provided herein are pharmaceutical compositions forinhalation administration containing: (i) an effective amount of adisclosed compound; optionally (ii) an effective amount of one or moresecond agents; and (iii) one or more pharmaceutical excipients suitablefor inhalation administration. In some embodiments, the pharmaceuticalcomposition further contains: (iv) an effective amount of a third agent.

Pharmaceutical compositions for inhalation or insufflation includesolutions and suspensions in pharmaceutically acceptable, aqueous ororganic solvents, or mixtures thereof, and powders. The liquid or solidpharmaceutical compositions can contain suitable pharmaceuticallyacceptable excipients as described herein. In some embodiments, thepharmaceutical compositions are administered by the oral or nasalrespiratory route for local or systemic effect. Pharmaceuticalcompositions in pharmaceutically acceptable solvents can be nebulized byuse of inert gases. Nebulized solutions can be inhaled directly from thenebulizing device or the nebulizing device can be attached to a facemask tent, or intermittent positive pressure breathing machine.Solution, suspension, or powder pharmaceutical compositions can beadministered, e.g., orally or nasally, from devices that deliver theformulation in an appropriate manner.

Also provided herein are formulations to be administered by inhalation.All types of inhalable formulation known in the art can be used inconnection with methods provided herein.

In a dry powder inhaler, the dose to be administered is stored in theform of a non-pressurized dry powder and, on actuation of the inhaler,the particles of the powder are inhaled by the patient. Dry powderinhalers can be “passive” devices in which the patient's breath is theonly source of gas which provides a motive force in the device, or“active” devices in which a source of compressed gas or alternativeenergy source is used. Formulations provided herein can be administeredwith either passive or active inhaler devices.

While it is desirable for as large a proportion as possible of theparticles of active material to be delivered to the deep lung, it isusually preferable for as little as possible of the other components topenetrate the deep lung. Therefore, powders generally include particlesof an active material, and carrier particles for carrying the particlesof active material. The carrier particles can be composed of anypharmacologically inert material or combination of materials which isacceptable for inhalation. In some embodiments, carrier particles arecomposed of one or more crystalline sugars. In some embodiments, thecarrier particles can be composed of one or more sugar alcohols orpolyols. In some embodiments, the carrier particles are particles ofdextrose or lactose. In some embodiments, the amount of carrierparticles is up to 95%, up to 90%, up to 80%, or up to 50% by weightbased on the total weight of the composition.

An additive material can also be provided in a dose which indicates tothe patient that the dose has been administered. (See, e.g., WO01/82906). The additive material, also referred to as indicatormaterial, can be present in the powder as formulated for the dry powderinhaler, or be present in a separate form, such as in a separatelocation within the inhaler such that the additive becomes entrained inthe airflow generated on inhalation simultaneously or sequentially withthe powder containing the active material. Accordingly, provided hereinis a formulation comprising a compound provided herein (e.g., Compound4), or an enantiomer, a mixture of enantiomers, or a mixture of two ormore diastereomers thereof, or a pharmaceutically acceptable formthereof, in combination with a carrier material.

Formulations provided herein, when inhaled, in some embodiments exhibita time to therapeutic effect of less than 3 hours, 2 hours, 1 hour, 30minutes, 15 minutes, 10 minutes, or 5 minutes. In some embodiments,formulations provided herein, when inhaled, will have a therapeuticduration of about 1 to 48 hours.

In certain embodiments of the present invention, each dose is stored ina “blister” of a blister pack. In this regard, since an active agent maybe susceptible to oxidation, it is sometimes important to prevent (orsubstantially limit) oxidation of the active agent prior toadministration. Thus, in some embodiments, exposure of the formulationto air prior to administration is prevented by storing each dose in asealed blister. In some embodiments, oxidation is further prevented (orlimited) by placing a plurality of blisters into a further sealedcontainer, such as a sealed bag made, for example of a foil such asaluminum foil. In some embodiments, the use of the sealed blisters (andoptional sealed bags) can minimize the need to include anti-oxidants inthe formulation.

In case of administration by a dry powder inhaler of the particles ofactive ingredient to the lung where they can be absorbed, the particlesize characteristics of the powder are particularly important. Inparticular, for the effective delivery of active ingredient deep intothe lung, the active particles should be small and well dispersed onactuation of the inhaler. In some embodiments, a fine particle fractionof at least 35% is generated on actuation of the inhaler device. In someembodiments, a fine particle fraction of at least 60%, at least 70%, orat least 80% is generated on actuation.

In certain embodiments, the formulation can also contain fine particlesof an excipient material, which can be a material such as one of thosereferred to above as being suitable for use as a carrier material, forexample, a crystalline sugar such as dextrose or lactose. The fineexcipient material can be of the same or a different material from thecarrier particles, where both are present. In certain embodiments, whereany carrier particles and/or any fine excipient material present is of amaterial itself capable of inducing a sensation in the oropharyngealregion, the carrier particles and/or the fine excipient material canalso be the indicator material. For example, the carrier particlesand/or any fine particle excipient can comprise mannitol. In certainembodiments, the amount of fine excipient material, if present, can beup to 50%, up to 30%, or up to 20%, by weight, based on the total weightof the composition.

Formulations provided herein can also be formulated with additionalexcipients to aid delivery and release. In certain embodiments, powdercan be formulated with relatively large carrier particles which aid theflow properties of the powder. Examples of large carrier particlesinclude, but are not limited to, lactose particles having a mass mediumaerodynamic diameter of greater than 90 microns. In some embodiments,hydrophobic microparticles can be dispersed within a carrier material.For example, the hydrophobic microparticles can be dispersed within apolysaccharide matrix, with the overall composition formulated asmicroparticles for direct delivery to the lung. The polysaccharide actsas a further barrier to the immediate release of the active agent. Thiscan further aid the controlled release process. An example of a suitablepolysaccharide is xanthan gum. Examples of hydrophobic materialsinclude, but are not limited to, solid state fatty acids such as oleicacid, lauric acid, palmitic acid, stearic acid, erucic acid, behenicacid, or derivatives (e.g., esters and salts) thereof. Specific examplesof such materials include, but are not limited to, phosphatidylcholines,phosphatidylglycerols and other natural and synthetic lung surfactants.In some embodiments, formulations provided herein contain metalstearates, in particular magnesium stearate, which has been approved fordelivery via the lung.

Formulations provided herein can also include one or more force controladditives (FCAs) in addition to the carrier and the active ingredient.In some embodiments, the FCAs can be provided in an amount from about0.1% to about 10% by weight, from about 0.15% to 5% by weight, or fromabout 0.5% to about 2% by weight of the total composition. In someembodiment, FCAs include, but are not limited to, anti-adherentmaterials. In some embodiments, FCAs include, but are not limited to,magnesium stearate, leucine, lecithin, and sodium stearyl fumarate, andthose described in U.S. Pat. No. 6,153,224, which is hereby incorporatedby reference.

In certain embodiments, formulations provided herein can be a “carrierfree” formulation, which includes only the active ingredient and one ormore anti-adherents. Such carrier free formulations are described in WO97/03649, the entire disclosure of which is hereby incorporated byreference.

As used herein, and unless otherwise specified, the term “anti-adherentmaterial” refers to those additive materials which will decrease thecohesion between the particles of the powder. Those materials willinclude, but are not limited to, leucine and lecithin. In someembodiments, the anti-adherent material comprises an amino acid. Aminoacids have been found to provide, when present as anti-adherentmaterial, high respirable fraction of the active material and also goodflow properties of the powder. In some embodiments, the amino acid isleucine, in particular L-leucine. In some embodiments, the D- andDL-forms can also be used. The anti-adherent material can comprise oneor more of any of the following amino acids: leucine, isoleucine,lysine, valine, methionine, cysteine, phenylalanine In some embodiments,the anti-adherent material can include magnesium stearate or colloidalsilicon dioxide.

In some embodiments, formulations provided herein are an aerosolformulation. In some embodiments, the aerosol formulation can becontained in a canister. Examples of aerosol formulation include, butare not limited to, an aerosol solution formulation and an aerosolsuspension formulation. In certain embodiments, the aerosol formulationcan contain a compound provided herein, optionally in combination withother active ingredient(s), in a propellant or in a propellant/solventsystem and, optionally, further pharmaceutical acceptable additive orexcipient.

The propellant can be any pressure-liquefied propellant and ispreferably a hydrofluoroalkane (HFA) or a mixture of different HFAs,including, but not limited to, HFA 134a (1,1,1,2-tetrafluoroethane), HFA227 (1,1,1,2,3,3,3-heptafluoropropane), and mixtures thereof.

The solvent generally has a higher polarity than that of the propellantand can include one or more substances such as a pharmaceuticallyacceptable alcohol (e.g., ethanol), a polyol, such as propylene glycolor polyethylene glycol, or mixtures thereof. In some embodiments, thesolvent is a lower branched or linear alkyl (C₁-C₄) alcohols such asethanol and isopropyl alcohol. In one embodiment, the co-solvent isethanol.

In some embodiments, the active ingredient of the formulation issubstantially completely and homogeneously dissolved in thepropellant/solvent system, i.e., the formulation is a solutionformulation.

Optionally, the formulation can comprise other pharmaceuticallyacceptable additives or excipients, which are substantially inertmaterials that are non-toxic and do not interact in negative manner withother components of the formulation. In some embodiments, theformulation can comprise one or more co-solvents, surfactants,carbohydrate, phospholipid, polymer, wetting agent, stabilizers,lubricants, or low volatility components.

In some embodiments, a suitable amount of an acid (organic or inorganicacid (mineral acids)) can be used as stabilizer. Examples include, butare not limited to, pharmaceutically acceptable monoprotic or polyproticacid, such as: hydrogen halides (hydrochloric acid, hydrobromic acid,hydroiodic acid, etc.), phosphoric acid, nitric acid, sulphuric acid,and halogen oxoacids.

In some embodiments, low volatility components can be used in order toincrease the mass median aerodynamic diameter (MMAD) of the aerosolparticles upon actuation of the inhaler and/or to improve the solubilityof the active ingredient in the propellant/solvent system. In someembodiments, the low volatility component has a vapor pressure at 25° C.lower than 0.1 kPa, or lower than 0.05 kPa. Examples of low-volatilitycomponents include, but are not limited to: esters such as isopropylmyristate, ascorbyl myristate, tocopherol esters; glycols such aspropylene glycol, polyethylene glycol, glycerol; and surface activeagents such as saturated organic carboxylic acids (e.g., lauric,myristic, stearic acid) and unsaturated carboxylic acids (e.g., oleic orascorbic acid). The amount of low volatility component can vary from 0.1to 10% w/w, from 0.5 to 5% (w/w), or from 1 to 2% (w/w).

In some embodiments, water at an amount between 0.005 and 0.3% (w/w) canbe added to the formulations in order to favorably affect the solubilityof the active ingredient without increasing the MMAD of the aerosoldroplets upon actuation.

1E. Formulations for Ocular Administration

In some embodiments, the disclosure provides a pharmaceuticalcomposition for treating ophthalmic disorders. The pharmaceuticalcomposition can contain an effective amount of a compound as disclosedherein and a pharmaceutical excipient suitable for ocularadministration. Pharmaceutical compositions suitable for ocularadministration can be presented as discrete dosage forms, such as dropsor sprays each containing a predetermined amount of an active ingredienta solution, or a suspension in an aqueous or non-aqueous liquid, anoil-in-water emulsion, or a water-in-oil liquid emulsion. Otheradministration forms include intraocular injection, intravitrealinjection, topically, or through the use of a drug eluting device,microcapsule, implant, or microfluidic device. In some cases, thecompounds as disclosed herein are administered with a carrier orexcipient that increases the intraocular penetrance of the compound suchas an oil and water emulsion with colloid particles having an oily coresurrounded by an interfacial film. It is contemplated that all localroutes to the eye can be used including topical, subconjunctival,periocular, retrobulbar, subtenon, intracameral, intravitreal,intraocular, subretinal, juxtascleral and suprachoroidal administration.Systemic or parenteral administration can be feasible including, but notlimited to intravenous, subcutaneous, and oral delivery. An exemplarymethod of administration will be intravitreal or subtenon injection ofsolutions or suspensions, or intravitreal or subtenon placement ofbioerodible or non-bioerodible devices, or by topical ocularadministration of solutions or suspensions, or posterior juxtascleraladministration of a gel or cream formulation.

Eye drops can be prepared by dissolving the active ingredient in asterile aqueous solution such as physiological saline, bufferingsolution, etc., or by combining powder compositions to be dissolvedbefore use. Other vehicles can be chosen, as is known in the art,including, but not limited to: balance salt solution, saline solution,water soluble polyethers such as polyethyene glycol, polyvinyls, such aspolyvinyl alcohol and povidone, cellulose derivatives such asmethylcellulose and hydroxypropyl methylcellulose, petroleum derivativessuch as mineral oil and white petrolatum, animal fats such as lanolin,polymers of acrylic acid such as carboxypolymethylene gel, vegetablefats such as peanut oil and polysaccharides such as dextrans, andglycosaminoglycans such as sodium hyaluronate. In some embodiments,additives ordinarily used in the eye drops can be added. Such additivesinclude isotonizing agents (e.g., sodium chloride, etc.), buffer agent(e.g., boric acid, sodium monohydrogen phosphate, sodium dihydrogenphosphate, etc.), preservatives (e.g., benzalkonium chloride,benzethonium chloride, chlorobutanol, etc.), thickeners (e.g.,saccharide such as lactose, mannitol, maltose, etc.; e.g., hyaluronicacid or its salt such as sodium hyaluronate, potassium hyaluronate,etc.; e.g., mucopolysaccharide such as chondroitin sulfate, etc.; e.g.,sodium polyacrylate, carboxyvinyl polymer, crosslinked polyacrylate,polyvinyl alcohol, polyvinyl pyrrolidone, methyl cellulose, hydroxypropyl methylcellulose, hydroxyethyl cellulose, carboxymethyl cellulose,hydroxy propyl cellulose or other agents known to those skilled in theart).

In some cases, the colloid particles include at least one cationic agentand at least one non-ionic surfactant such as a poloxamer, tyloxapol, apolysorbate, a polyoxyethylene castor oil derivative, a sorbitan ester,or a polyoxyl stearate. In some cases, the cationic agent is analkylamine, a tertiary alkyl amine, a quarternary ammonium compound, acationic lipid, an amino alcohol, a biguanidine salt, a cationiccompound or a mixture thereof. In some cases, the cationic agent is abiguanidine salt such as chlorhexidine, polyaminopropyl biguanidine,phenformin, alkylbiguanidine, or a mixture thereof. In some cases, thequaternary ammonium compound is a benzalkonium halide, lauralkoniumhalide, cetrimide, hexadecyltrimethylammonium halide,tetradecyltrimethylammonium halide, dodecyltrimethylammonium halide,cetrimonium halide, benzethonium halide, behenalkonium halide,cetalkonium halide, cetethyldimonium halide, cetylpyridinium halide,benzododecinium halide, chlorallyl methenamine halide, rnyristylalkoniumhalide, stearalkonium halide or a mixture of two or more thereof. Insome cases, cationic agent is a benzalkonium chloride, lauralkoniumchloride, benzododecinium bromide, benzethenium chloride,hexadecyltrimethylammonium bromide, tetradecyltrimethylammonium bromide,dodecyltrimethylammonium bromide or a mixture of two or more thereof. Insome cases, the oil phase is mineral oil and light mineral oil, mediumchain triglycerides (MCT), coconut oil; hydrogenated oils comprisinghydrogenated cottonseed oil, hydrogenated palm oil, hydrogenate castoroil or hydrogenated soybean oil; polyoxyethylene hydrogenated castor oilderivatives comprising poluoxyl-40 hydrogenated castor oil, polyoxyl-60hydrogenated castor oil or polyoxyl-100 hydrogenated castor oil.

1F. Formulations for Controlled Release Administration

In some embodiments, provided herein are pharmaceutical compositions forcontrolled release administration containing a compound as disclosedherein, and a pharmaceutical excipient suitable for controlled releaseadministration. In some embodiments, provided herein are pharmaceuticalcompositions for controlled release administration containing: (i) aneffective amount of a disclosed compound; optionally (ii) an effectiveamount of one or more second agents; and (iii) one or morepharmaceutical excipients suitable for controlled releaseadministration. In some embodiments, the pharmaceutical compositionfurther contains: (iv) an effective amount of a third agent.

Active agents such as the compounds provided herein can be administeredby controlled release means or by delivery devices that are well knownto those of ordinary skill in the art. Examples include, but are notlimited to, those described in U.S. Pat. Nos. 3,845,770; 3,916,899;3,536,809; 3,598,123; and 4,008,719; 5,674,533; 5,059,595; 5,591,767;5,120,548; 5,073,543; 5,639,476; 5,354,556; 5,639,480; 5,733,566;5,739,108; 5,891,474; 5,922,356; 5,972,891; 5,980,945; 5,993,855;6,045,830; 6,087,324; 6,113,943; 6,197,350; 6,248,363; 6,264,970;6,267,981; 6,376,461; 6,419,961; 6,589,548; 6,613,358; 6,699,500 each ofwhich is incorporated herein by reference. Such dosage forms can be usedto provide slow or controlled release of one or more active agentsusing, for example, hydropropylmethyl cellulose, other polymer matrices,gels, permeable membranes, osmotic systems, multilayer coatings,microparticles, liposomes, microspheres, or a combination thereof toprovide the desired release profile in varying proportions. Suitablecontrolled release formulations known to those of ordinary skill in theart, including those described herein, can be readily selected for usewith the active agents provided herein. Thus, the pharmaceuticalcompositions provided encompass single unit dosage forms suitable fororal administration such as, but not limited to, tablets, capsules,gelcaps, and caplets that are adapted for controlled release.

All controlled release pharmaceutical products have a common goal ofimproving drug therapy over that achieved by their non controlledcounterparts. In some embodiments, the use of a controlled releasepreparation in medical treatment is characterized by a minimum of drugsubstance being employed to cure or control the disease, disorder, orcondition in a minimum amount of time. Advantages of controlled releaseformulations include extended activity of the drug, reduced dosagefrequency, and increased subject compliance. In addition, controlledrelease formulations can be used to affect the time of onset of actionor other characteristics, such as blood levels of the drug, and can thusaffect the occurrence of side (e.g., adverse) effects.

In some embodiments, controlled release formulations are designed toinitially release an amount of a compound as disclosed herein thatpromptly produces the desired therapeutic effect, and gradually andcontinually release other amounts of the compound to maintain this levelof therapeutic or prophylactic effect over an extended period of time.In order to maintain this constant level of the compound in the body,the compound should be released from the dosage form at a rate that willreplace the amount of drug being metabolized and excreted from the body.Controlled release of an active agent can be stimulated by variousconditions including, but not limited to, pH, temperature, enzymes,water, or other physiological conditions or compounds.

In certain embodiments, the pharmaceutical composition can beadministered using intravenous infusion, an implantable osmotic pump, atransdermal patch, liposomes, or other modes of administration. In oneembodiment, a pump can be used (see, Sefton, CRC Crit. Ref. Biomed. Eng.14:201 (1987); Buchwald et al., Surgery 88:507 (1980); Saudek et al., N.Engl. J. Med. 321:574 (1989)). In another embodiment, polymericmaterials can be used. In yet another embodiment, a controlled releasesystem can be placed in a subject at an appropriate site determined by apractitioner of skill, e.g., thus requiring only a fraction of thesystemic dose (see, e.g., Goodson, Medical Applications of ControlledRelease, 115-138 (vol. 2, 1984). Other controlled release systems arediscussed in the review by Langer, Science 249:1527-1533 (1990). The oneor more active agents can be dispersed in a solid inner matrix, e.g.,polymethylmethacrylate, polybutylmethacrylate, plasticized orunplasticized polyvinylchloride, plasticized nylon, plasticizedpolyethyleneterephthalate, natural rubber, polyisoprene,polyisobutylene, polybutadiene, polyethylene, ethylene-vinylacetatecopolymers, silicone rubbers, polydimethylsiloxanes, silicone carbonatecopolymers, hydrophilic polymers such as hydrogels of esters of acrylicand methacrylic acid, collagen, cross-linked polyvinylalcohol andcross-linked partially hydrolyzed polyvinyl acetate, that is surroundedby an outer polymeric membrane, e.g., polyethylene, polypropylene,ethylene/propylene copolymers, ethylene/ethyl acrylate copolymers,ethylene/vinylacetate copolymers, silicone rubbers, polydimethylsiloxanes, neoprene rubber, chlorinated polyethylene, polyvinylchloride,vinylchloride copolymers with vinyl acetate, vinylidene chloride,ethylene and propylene, ionomer polyethylene terephthalate, butyl rubberepichlorohydrin rubbers, ethylene/vinyl alcohol copolymer,ethylene/vinyl acetate/vinyl alcohol terpolymer, andethylene/vinyloxyethanol copolymer, that is insoluble in body fluids.The one or more active agents then diffuse through the outer polymericmembrane in a release rate controlling step. The percentage of activeagent in such parenteral compositions is highly dependent on thespecific nature thereof, as well as the needs of the subject.

Dosage

A compound described herein (e.g., a PI3K-γ inhibitor such as Compound4) can be delivered in the form of pharmaceutically acceptablecompositions which comprise a therapeutically effective amount of one ormore compounds described herein and/or one or more additionaltherapeutic agents such as a chemotherapeutic, formulated together withone or more pharmaceutically acceptable excipients. In some instances,the compound described herein and the additional therapeutic agent areadministered in separate pharmaceutical compositions and can (e.g.,because of different physical and/or chemical characteristics) beadministered by different routes (e.g., one therapeutic is administeredorally, while the other is administered intravenously). In otherinstances, the compound described herein and the additional therapeuticagent can be administered separately, but via the same route (e.g., bothorally or both intravenously). In still other instances, the compounddescribed herein and the additional therapeutic agent can beadministered in the same pharmaceutical composition.

The selected dosage level will depend upon a variety of factorsincluding, for example, the activity of the particular compoundemployed, the route of administration, the time of administration, therate of excretion or metabolism of the particular compound beingemployed, the rate and extent of absorption, the duration of thetreatment, other drugs, compounds and/or materials used in combinationwith the particular compound employed, the age, sex, weight, condition,general health and prior medical history of the patient being treated,and like factors well known in the medical arts.

In general, a suitable daily dose of a compound described herein and/ora chemotherapeutic will be that amount of the compound which, in someembodiments, can be the lowest dose effective to produce a therapeuticeffect. Such an effective dose will generally depend upon the factorsdescribed herein. Generally, doses of the compounds described herein fora patient, when used for the indicated effects, will range from about0.0001 mg to about 100 mg per day, or about 0.001 mg to about 100 mg perday, or about 0.01 mg to about 100 mg per day, or about 0.1 mg to about100 mg per day, or about 0.0001 mg to about 500 mg per day, or about0.001 mg to about 500 mg per day, or about 0.01 mg to 1000 mg, or about0.01 mg to about 500 mg per day, or about 0.1 mg to about 500 mg perday, or about 1 mg to 50 mg per day, or about 5 mg to 40 mg per day. Insome embodiments, range is from about 1 mg to about 100 mg, about 1 mgto about 200 mg, about 1 mg to about 500 mg, about 1 mg to about 1000mg, about 100 mg to about 200 mg, about 100 mg to about 500 mg, about100 to about 750 mg, about 100 mg to about 1000 mg. An exemplary dosageis about 10 to 30 mg per day. In some embodiments, for a 70 kg human, asuitable dose would be about 0.05 to about 7 g/day, such as about 0.05to about 2.5 g/day. Actual dosage levels of the active ingredients inthe pharmaceutical compositions described herein can be varied so as toobtain an amount of the active ingredient which is effective to achievethe desired therapeutic response for a particular patient, composition,and mode of administration, without being toxic to the patient. In someinstances, dosage levels below the lower limit of the aforesaid rangecan be more than adequate, while in other cases still larger doses canbe employed without causing any harmful side effect, e.g., by dividingsuch larger doses into several small doses for administration throughoutthe day.

Pharmacokinetic studies in animals provided herein suggest efficaciousdose ranges for Compound 4. Generally speaking, to specifically inhibitPI3K-γ, one can administer a dose of a specific PI3K-γ inhibitor thatresults in an unbound plasma concentration of the drug that is above apredetermined threshold (e.g., the IC50, IC60, IC70, IC80, or IC90 forPI3K-γ) for a selected time (e.g., 1 hour, 2 h, 3 h, 6 h, 12 h, 24 h, 2days, 3 d, 5 d, or 7 d). This dose may be selected such that the plasmaconcentration is below a second predetermined threshold (e.g., the IC20,IC30, IC40, or IC50 for PI3K-δ, -α, or β) the for a selected time (e.g.,1 hour, 2 h, 3 h, 6 h, 12 h, 24 h, 2 days, 3 d, 5 d, or 7 d). In someembodiments, the PI3K-γ inhibitor, e.g., Compound 4, is administered ata dose that results in an unbound plasma concentration of Compound 4that is above the IC90 of PI3K-γ for at least 1 hour, 2 h, 3 h, 6 h, 12h, or 24 h. In some embodiments, the PI3K-γ inhibitor, e.g., Compound 4,is administered at a dose that results in an unbound plasmaconcentration of Compound 4 that is above the IC50 of PI3K-γ for atleast 1 hour, 2 h, 3 h, 6 h, 12 h, or 24 h.

Based on non-human animal studies, a predicted human dose to achieveexposure at the IC90 for PI3K-γ is approximately 2 mg. Accordingly, insome embodiments, the methods herein involve administering a selectivePI3K-γ inhibitor, e.g., Compound 4, to a human, wherein each dose isabout 2 mg, 1-3 mg, 1-5 mg, 1-10 mg, 0.5-20 mg, or 0.1-50 mg. In someembodiments, the dose (e.g., a therapeutically effective dose) is about2 mg, 1-3 mg, 1-5 mg, 1-10 mg, 0.5-20 mg, 0.1-50 mg, 0.1-75 mg, 0.5-75mg, 1-75 mg, 0.1-100 mg, 0.5-100 mg, or 1-100 mg. In some embodiments,the dose is about 1-10 mg. In some embodiments, the dose is about 1-50mg. In some embodiments, the dose is about 1-100 mg. In a 70 kg human, a2 mg dose corresponds to 0.029 mg/kg. Accordingly, in some embodiments,the methods herein involve administering a selective PI3K-γ inhibitor,e.g., Compound 4, to a human, wherein each dose is about 0.029 mg/kg,0.014-0.14 mg/kg, 0.02-0.04 mg/kg, 0.01-0.05 mg/kg, 0.01-0.1, or0.01-0.5 mg/kg.

In some embodiments, the compounds can be administered daily, everyother day, three times a week, twice a week, weekly, or bi-weekly. Thedosing schedule can include a “drug holiday,” e.g., the drug can beadministered for two weeks on, one week off, or three weeks on, one weekoff, or four weeks on, one week off, etc., or continuously, without adrug holiday. The compounds can be administered orally, intravenously,intraperitoneally, topically, transdermally, intramuscularly,subcutaneously, intranasally, sublingually, or by any other route.

In some embodiments, a compound as provided herein is administered inmultiple doses. Dosing can be about once, twice, three times, fourtimes, five times, six times, or more than six times per day. Dosing canbe about once a month, about once every two weeks, about once a week, orabout once every other day. In another embodiment, a compound asdisclosed herein and another agent are administered together from aboutonce per day to about 6 times per day. In another embodiment, theadministration of a compound as provided herein and an agent continuesfor less than about 7 days. In yet another embodiment, theadministration continues for more than about 6 days, about 10 days,about 14 days, about 28 days, about two months, about six months, orabout one year. In some cases, continuous dosing is achieved andmaintained as long as necessary.

Based on non-human animal studies provided herein the oral half-life ofCompound 4 in humans is expected to be about 10-13 hours. This findinginforms the timing of administration of a PI3K-γ inhibitor such asCompound 4. For instance, in some embodiments, the timing is selectedsuch that an unbound plasma concentration of the drug that is above apredetermined threshold (e.g., the IC50, IC60, IC70, IC80, or IC90 forPI3K-γ) for a selected time (e.g., 1 hour, 2 h, 3 h, 6 h, 12 h, 24 h, 2days, 3 d, 5 d, or 7 d). The timing of administration may also be chosensuch that the plasma level is below a second predetermined threshold(e.g., the IC20, IC30, IC40, or IC50 for PI3K-δ, -α, or β) the for aselected time (e.g., 1 hour, 2 h, 3 h, 6 h, 12 h, 24 h, 2 days, 3 d, 5d, or 7 d). In some embodiments, the PI3K-γ inhibitor, e.g., Compound 4,is administered with timing that results in an unbound plasmaconcentration of Compound 4 that is above the IC90 of PI3K-γ for atleast 1 hour, 2 h, 3 h, 6 h, 12 h, or 24 h. In some embodiments, thePI3K-γ inhibitor, e.g., Compound 4, is administered with timing thatresults in an unbound plasma concentration of Compound 4 that is abovethe IC50 of PI3K-γ for at least 1 hour, 2 h, 3 h, 6 h, 12 h, or 24 h.

Accordingly, in some embodiments, the methods herein involveadministering a selective PI3K-γ inhibitor, e.g., Compound 4, to ahuman, about once per day. In embodiments, the selective PI3K-γinhibitor, e.g., Compound 4, is administered to a human once every twodays. In embodiments, the selective PI3K-γ inhibitor, e.g., Compound 4,is administered to a human twice or three times per day.

Administration of the pharmaceutical compositions as disclosed hereincan continue as long as necessary. In some embodiments, an agent asdisclosed herein is administered for more than about 1, about 2, about3, about 4, about 5, about 6, about 7, about 14, or about 28 days. Insome embodiments, an agent as disclosed herein is administered for lessthan about 28, about 14, about 7, about 6, about 5, about 4, about 3,about 2, or about 1 day. In some embodiments, an agent as disclosedherein is administered chronically on an ongoing basis, e.g., for thetreatment of chronic effects.

Since the compounds described herein can be administered in combinationwith other treatments (such as additional chemotherapeutics, radiationor surgery), the doses of each agent or therapy can be lower than thecorresponding dose for single-agent therapy. The dose for single-agenttherapy can range from, for example, about 0.0001 to about 200 mg, orabout 0.001 to about 100 mg, or about 0.01 to about 100 mg, or about 0.1to about 100 mg, or about 1 to about 50 mg per kilogram of body weightper day. In some embodiments, the dose is about 1 mg/kg, about 5 mg/kg,about 7.5 mg/kg, about 10 mg/kg, about 15 mg/kg, about 20 mg/kg, about25 mg/kg, about 50 mg/kg, about 75 mg/kg, or about 100 mg/kg per day. Insome embodiments, the dose is about 1 mg/kg, about 7.5 mg/kg, about 20mg/kg, or about 50 mg/kg per day.

When a compound provided herein, is administered in a pharmaceuticalcomposition that comprises one or more agents, and the agent has ashorter half-life than the compound provided herein unit dose forms ofthe agent and the compound provided herein can be adjusted accordingly.

Kits

In some embodiments, provided herein are kits. The kits can include acompound or pharmaceutical composition as described herein, in suitablepackaging, and written material that can include instructions for use,discussion of clinical studies, listing of side effects, and the like.Such kits can also include information, such as scientific literaturereferences, package insert materials, clinical trial results, and/orsummaries of these and the like, which indicate or establish theactivities and/or advantages of the pharmaceutical composition, and/orwhich describe dosing, administration, side effects, drug interactions,or other information useful to the health care provider. Suchinformation can be based on the results of various studies, for example,studies using experimental animals involving in vivo models and studiesbased on human clinical trials.

In some embodiments, a memory aid is provided with the kit, e.g., in theform of numbers next to the tablets or capsules whereby the numberscorrespond with the days of the regimen which the tablets or capsules sospecified should be ingested. Another example of such a memory aid is acalendar printed on the card, e.g., as follows “First Week, Monday,Tuesday, . . . etc. . . . Second Week, Monday, Tuesday, . . . ” etc.Other variations of memory aids will be readily apparent. A “daily dose”can be a single tablet or capsule or several tablets or capsules to betaken on a given day.

The kit can further contain another agent. In some embodiments, thecompound as disclosed herein and the agent are provided as separatepharmaceutical compositions in separate containers within the kit. Insome embodiments, the compound as disclosed herein and the agent areprovided as a single pharmaceutical composition within a container inthe kit. Suitable packaging and additional articles for use (e.g.,measuring cup for liquid preparations, foil wrapping to minimizeexposure to air, and the like) are known in the art and can be includedin the kit. In other embodiments, kits can further comprise devices thatare used to administer the active agents. Examples of such devicesinclude, but are not limited to, syringes, drip bags, patches, andinhalers. Kits described herein can be provided, marketed and/orpromoted to health providers, including physicians, nurses, pharmacists,formulary officials, and the like. Kits can also, in some embodiments,be marketed directly to the consumer.

An example of such a kit is a so-called blister pack. Blister packs arewell known in the packaging industry and are being widely used for thepackaging of pharmaceutical unit dosage forms (tablets, capsules, andthe like). Blister packs generally consist of a sheet of relativelystiff material covered with a foil of a preferably transparent plasticmaterial. During the packaging process, recesses are formed in theplastic foil. The recesses have the size and shape of the tablets orcapsules to be packed. Next, the tablets or capsules are placed in therecesses and the sheet of relatively stiff material is sealed againstthe plastic foil at the face of the foil which is opposite from thedirection in which the recesses were formed. As a result, the tablets orcapsules are sealed in the recesses between the plastic foil and thesheet. The strength of the sheet is such that the tablets or capsulescan be removed from the blister pack by manually applying pressure onthe recesses whereby an opening is formed in the sheet at the place ofthe recess. The tablet or capsule can then be removed via said opening.

Kits can further comprise pharmaceutically acceptable vehicles that canbe used to administer one or more active agents. For example, if anactive agent is provided in a solid form that must be reconstituted forparenteral administration, the kit can comprise a sealed container of asuitable vehicle in which the active agent can be dissolved to form aparticulate-free sterile solution that is suitable for parenteraladministration. Examples of pharmaceutically acceptable vehiclesinclude, but are not limited to: Water for Injection USP; aqueousvehicles such as, but not limited to, Sodium Chloride Injection,Ringer's Injection, Dextrose Injection, Dextrose and Sodium ChlorideInjection, and Lactated Ringer's Injection; water-miscible vehicles suchas, but not limited to, ethyl alcohol, polyethylene glycol, andpolypropylene glycol; and non-aqueous vehicles such as, but not limitedto, corn oil, cottonseed oil, peanut oil, sesame oil, ethyl oleate,isopropyl myristate, and benzyl benzoate.

The present disclosure further encompasses anhydrous pharmaceuticalcompositions and dosage forms comprising an active ingredient, sincewater can facilitate the degradation of some compounds. For example,water can be added (e.g., about 5%) in the pharmaceutical arts as ameans of simulating long-term storage in order to determinecharacteristics such as shelf-life or the stability of formulations overtime. Anhydrous pharmaceutical compositions and dosage forms can beprepared using anhydrous or low moisture containing ingredients and lowmoisture or low humidity conditions. For example, pharmaceuticalcompositions and dosage forms which contain lactose can be madeanhydrous if substantial contact with moisture and/or humidity duringmanufacturing, packaging, and/or storage is expected. An anhydrouspharmaceutical composition can be prepared and stored such that itsanhydrous nature is maintained. Accordingly, anhydrous pharmaceuticalcompositions can be packaged using materials known to prevent exposureto water such that they can be included in suitable formulary kits.Examples of suitable packaging include, but are not limited to,hermetically sealed foils, plastic or the like, unit dose containers,blister packs, and strip packs.

Therapeutic Methods

Phosphoinositide 3-kinases (PI3Ks) are members of a conserved family oflipid kinases that regulate numerous cell functions, includingproliferation, differentiation, cell survival and metabolism. Severalclasses of PI3Ks exist in mammalian cells, including Class IA subgroup(e.g., PI3K-α, β, δ), which are generally activated by receptor tyrosinekinases (RTKs); Class IB (e.g., PI3K-γ), which is activated by G-proteincoupled receptors (GPCRs), among others. PI3Ks exert their biologicalactivities via a “PI3K-mediated signaling pathway” that includes severalcomponents that directly and/or indirectly transduce a signal triggeredby a PI3K, including the generation of second messengerphophotidylinositol, 3,4,5-triphosphate (PIP3) at the plasma membrane,activation of heterotrimeric G protein signaling, and generation offurther second messengers such as cAMP, DAG, and IP3, all of which leadsto an extensive cascade of protein kinase activation (reviewed inVanhaesebroeck, B. et al. (2001) Annu Rev Biochem. 70:535-602). Forexample, PI3K-δ is activated by cellular receptors through interactionbetween the PI3K regulatory subunit (p85) SH2 domains, or through directinteraction with RAS. PIP3 produced by PI3K activates effector pathwaysdownstream through interaction with plextrin homology (PH) domaincontaining enzymes (e.g., PDK-1 and AKT [PKB]). (Fung-Leung W P. (2011)Cell Signal. 23(4):603-8). Unlike PI3K-δ, PI3K-γ is not associated witha regulatory subunit of the p85 family, but rather with a regulatorysubunit in the p101 or p84 families. PI3K-γ is associated with GPCRs,and is responsible for the very rapid induction of PIP3. PI3K-γ can bealso activated by RAS.

In some embodiments, provided herein are methods of modulating a PI3kinase activity (e.g., selectively modulating) by contacting the kinasewith an effective amount of a compound as provided herein, or apharmaceutically acceptable form (e.g., pharmaceutically acceptablesalts, hydrates, solvates, isomers, prodrugs, and isotopically labeledderivatives) thereof, or a pharmaceutical composition as providedherein. Modulation can be inhibition (e.g., reduction) or activation(e.g., enhancement) of kinase activity. In some embodiments, providedherein are methods of inhibiting kinase activity by contacting thekinase with an effective amount of a compound as provided herein insolution. In some embodiments, provided herein are methods of inhibitingthe kinase activity by contacting a cell, tissue, organ that express thekinase of interest, with a compound provided herein. In someembodiments, provided herein are methods of inhibiting kinase activityin a subject by administering into the subject an effective amount of acompound as provided herein, or a pharmaceutically acceptable formthereof. In some embodiments, the kinase activity is inhibited (e.g.,reduced) by more than about 25%, 30%, 40%, 50%, 60%, 70%, 80%, or 90%,when contacted with a compound provided herein as compared to the kinaseactivity without such contact. In some embodiments, provided herein aremethods of inhibiting PI3 kinase activity in a subject (includingmammals such as humans) by contacting said subject with an amount of acompound as provided herein sufficient to inhibit or reduce the activityof the PI3 kinase in said subject.

In some embodiments, the kinase is a lipid kinase or a protein kinase.In some embodiments, the kinase is selected from a PI3 kinase includingdifferent isoforms, such as PI3 kinase α, PI3 kinase β, PI3 kinase γ,PI3 kinase δ; DNA-PK; mTOR; Abl, VEGFR, Ephrin receptor B4 (EphB4); TEKreceptor tyrosine kinase (TIE2); FMS-related tyrosine kinase 3 (FLT-3);Platelet derived growth factor receptor (PDGFR); RET; ATM; ATR; hSmg-1;Hck; Src; Epidermal growth factor receptor (EGFR); KIT; Insulin Receptor(IR); and IGFR.

As used herein, a “PI3K-mediated disorder” refers to a disease orcondition involving aberrant PI3K-mediated signaling pathway. In oneembodiment, provided herein is a method of treating a PI3K mediateddisorder in a subject, the method comprising administering atherapeutically effective amount of a compound as provided herein, or apharmaceutically acceptable form thereof, or a pharmaceuticalcomposition as provided herein. In some embodiments, provided herein isa method of treating a PI3K-δ or PI3K-γ mediated disorder in a subject,the method comprising administering a therapeutically effective amountof a compound as provided herein, or a pharmaceutically acceptable formthereof, or a pharmaceutical composition as provided herein. In someembodiments, provided herein is a method for inhibiting at least one ofPI3K-δ and PI3K-γ, the method comprising contacting a cell expressingPI3K in vitro or in vivo with an effective amount of a compound orcomposition provided herein. PI3Ks have been associated with a widerange of conditions, including immunity, cancer and thrombosis (reviewedin Vanhaesebroeck, B. et al. (2010) Current Topics in Microbiology andImmunology, DOI 10.1007/82_2010_65). For example, Class I PI3Ks,particularly PI3K-γ and PI3K-δ isoforms, are highly expressed inleukocytes and have been associated with adaptive and innate immunity;thus, these PI3Ks are believed to be important mediators in inflammatorydisorders and hematologic malignancies (reviewed in Harris, S J et al.(2009) Curr Opin Investig Drugs 10(11):1151-62); Rommel C. et al. (2007)Nat Rev Immunol 7(3):191-201; Durand C A et al. (2009) J Immunol.183(9):5673-84; Dil N, Marshall A J. (2009) Mol Immunol. 46(10):1970-8;Al-Alwan M M et al. (2007) J Immunol. 178(4):2328-35; Zhang T T, et al.(2008) J Allergy Clin Immunol. 2008; 122(4):811-819.e2; Srinivasan L, etal. (2009) Cell 139(3):573-86)

PI3K-γ Activities

PI3K-γ is a Class 1B PI3K that associates with the p101 and p84(p87PIKAP) adaptor proteins, and canonically signals through GPCRs.Non-canonical activation through tyrosine kinase receptors and RAS canoccur. Activated PI3K-γ leads to production of PIP3, which serves as adocking site for downstream effector proteins including AKT and BTK,bringing these enzymes to the cell membrane where they may be activated.A scaffolding role for PI3K-γ has been proposed and may contribute tothe activation of the RAS/MEK/ERK pathway. The interaction with the RASpathway explains activities attributed to kinase dead PI3K-γ in cells orin animals. PI3K-γ is essential for function of a variety of immunecells and pathways. Chemokine responses (including IL-8, fMLP, and C5a),leading to neutrophil, basophil or monocyte cell migration, is dependenton PI3K-γ (HIRSCH et al., “Central Role for G Protein-CoupledPhosphoinositide 3-Kinase γ in Inflammation,” Science 287:1049-1053(2000); SASAKI et al., “Function of PI3Kγ in Thymocyte Development, TCell Activation, and Neutrophil Migration,” Science 287:1040-1046(2000); L I et al., “Roles of PLC-β2 and -β3 and PI3Kγ inChemoattractant-Mediated Signal Transduction,” Science 287:1046-1049(2000)). The requirement for PI3K-γ-dependent neutrophil migration isdemonstrated by failure of arthritis development in the K/BXN serumtransfer arthritis model in PI3K-γ knockout mice (Randis et al., Eur. J.Immunol., 2008, 38(5), 1215-24). Similarly, the mice fail to developcellular inflammation and airway hyper-responsiveness in the ovalbumininduced asthma model (Takeda et al., J. Allergy Clin. Immunol., 2009;123, 805-12). PI3K-γ deficient mice also have defects in T-helper cellfunction. T-cell cytokine production and proliferation in response toactivation is reduced, and T helper dependent viral clearance isdefective (Sasaki et al., Science, 2000, 287, 1040-46). T cell dependentinflammatory disease models including EAE also do not develop in PI3K-γdeficient mice, and both the T-cell activation defect and cellularmigration defects may contribute to efficacy in this model (Comerfold,PLOS One, 2012, 7, e45095). The imiquimod psoriasis model has also beenused to demonstrate the importance of PI3K-γ in the inflammatoryresponse. Using PI3K-γ deficient mice in this model, the accumulation ofγδ T cells in the skin is blocked, as well as dendritic cell maturationand migration (ROLLER et al., “Blockade of Phosphatidylinositol 3-Kinase(PI3K)δ or PI3Kγ Reduces IL-17 and Ameliorates Imiquimod-InducedPsoriasis-like Dermatitis,” J. Immunol. 189:4612-4620 (2012)). The roleof PI3K-γ in cellular trafficking can also be demonstrated in oncologymodels where tumor inflammation is important for growth and metastasisof cancers. In the Lewis Lung Carcinoma model, monocyte activation,migration, and differentiation in tumors are defective. This defectresults in a reduction in tumor growth and extended survival in PI3K-γdeficient mice (Schmid et al., Cancer Cell, 2011, 19, 715-27) or upontreatment with inhibitors that target PI3K-γ. In pancreatic cancer,PI3K-γ can be inappropriately expressed, and in this solid tumor canceror others where PI3K-γ plays a functional role, inhibition of PI3K-γ canbe beneficial.

For instance, while not wishing to be bound by theory, PI3K-γ isexpressed in Gr1+CD11b+ myeloid cells, and directly promotes myeloidcell invasion and consequently, immunosuppression of pancreatic ductalcarcinomas. Hardamon et. al., Proceedings: AACR 103rd Annual Meeting2012, Cancer Research: Apr. 15, 2012; Volume 72, Issue 8, Supplement 1.Inhibition of PI3K-γ also shows promise for the treatment of hematologicmalignancies. In a T-ALL model employing a T cell directed knockout ofpten, PI3K-δ and PI3K-γ are both essential for the appropriatedevelopment of disease, as shown with genetic deletion of both genes(Subramaniam et al. Cancer Cell 21, 459-472, 2012). In addition, in thisT-ALL model, treatment with a small molecule inhibitor of both kinasesleads to extended survival of these mice. In CLL, chemokine networkssupport a pseudo-follicular microenvironment that includes Nurse likecells, stromal cells and T-helper cells. The roles of PI3K-γ in thenormal chemokine signaling and T cell biology suggest the value ofinhibiting this target in CLL (BURGER, “Inhibiting B-Cell ReceptorSignaling Pathways in Chronic Lymphocytic Leukemia,” Curr. Mematol.Malig. Rep. 7:26-33 (2012)). Accordingly, PI3K-γ inhibitors aretherapeutically interesting for diseases of the immune system where celltrafficking and T cell or myeloid cell function is important. Inoncology, solid tumors that are dependent on tumor inflammation, ortumors with high levels of PI3K-γ expression, can be targeted. Forhematological cancers, a special role for PI3K-γ and PI3K-δ isoforms inTALL and potentially in CLL suggests targeting these PI3Ks in thesediseases.

Without being limited by a particular theory, PI3K-γ has been shown toplay roles in inflammation, arthritis, asthma, allergy, multiplesclerosis (MS), and cancer, among others (e.g., Ruckle et al., NatureRev., Drug Discovery, 2006, 5, 903-18; Schmid et al., “Myeloid cells intumor inflammation,” Vascular Cell, 2012, doi:10.1186/2045-824X-4-14).For example, PI3K-γ functions in multiple signaling pathways involved inleukocyte activation and migration. PI3K-γ has been shown to drivepriming and survival of autoreactive CD4⁺ T cells during experimentalautoimmune encephalomyelitis (EAE), a model for MS. When administeredfrom onset of EAE, a PI3K-γ inhibitor has been shown to cause inhibitionand reversal of clinical disease, and reduction of demyelination andcellular pathology in the CNS (Comerford et al., PLOS One, 2012, 7,e45095). PI3K-γ also regulates thymocyte development, T cell activation,neutrophil migration, and the oxidative burst (Sasaki et al., Science,2000, 287, 1040-46). In addition, it is shown that allergic airwayhyper-responsiveness, inflammation, and remodeling do not develop inPI3K-γ deficient mice (Takeda et al., J. Allergy Clin. Immunol., 2009;123, 805-12). PI3K-γ is shown to be required for chemoattractant-inducedproduction of phosphatidylinositol 3,4,5-trisphosphate and has animportant role in chemoattractant-induced superoxide production andchemotaxis in mouse neutrophils and in production of T cell-independentantigen-specific antibodies composed of the immunoglobulin light chain(Li et al., Science, 2000, 287, 1046-49). PI3K-γ is reported to be acrucial signaling molecule required for macrophage accumulation ininflammation (Hirsch et al., Science, 2000, 287, 1049-53). In cancers,pharmacological or genetic blockade of p110γ suppresses inflammation,growth, and metastasis of implanted and spontaneous tumors, suggestingthat PI3K-γ can be an important therapeutic target in oncology (Schmidet al., Cancer Cell, 2011, 19, 715-27). For example, it is shown thatPI3K-γ has a tumor-specific high accumulation in pancreatic ductaladenocarcinoma (PDAC) in human, signifying a role of PI3K-γ inpancreatic cancer (Edling et al., Human Cancer Biology, 2010, 16(2),4928-37).

In certain embodiments, provided herein are methods of treating orpreventing a PI3K-gamma mediated disorder in a subject, comprisingadministering to the subject a therapeutically effective amount of acompound provided herein (e.g., a compound of any of Formulae (I″),(I′), (A′), (I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X),(XI), (XII), (XIII), (XIV), (XV), (XVI), and (XVII), e.g., a selectivePI3K-γ inhibitor, e.g., Compound 4), or a pharmaceutically acceptableform thereof.

In one embodiment, the subject has or is at risk of having a PI3K-gammamediated disorder selected from cancer, an inflammatory disease, or anautoimmune disease. In one embodiment, the cancer is a solid tumor. Inone embodiment, the cancer is selected from one or more of: a cancer ofthe pulmonary system, a brain cancer, a cancer of the gastrointestinaltract, a skin cancer, a genitourinary cancer, a pancreatic cancer, alung cancer, a medullobastoma, a basal cell carcinoma, a glioma, abreast cancer, a prostate cancer, a testicular cancer, an esophagealcancer, a hepatocellular cancer, a gastric cancer, a gastrointestinalstromal tumor (GIST), a colon cancer, a colorectal cancer, an ovariancancer, a melanoma, a neuroectodermal tumor, head and neck cancer, asarcoma, a soft-tissue sarcoma, fibrosarcoma, myxosarcoma, liposarcoma,a chondrosarcoma, an osteogenic sarcoma, a chordoma, an angiosarcoma, anendotheliosarcoma, a lymphangiosarcoma, a lymphangioendotheliosarcoma, asynovioma, a mesothelioma, a leiomyosarcoma, a cervical cancer, auterine cancer, an endometrial cancer, a carcinoma, a bladder carcinoma,an epithelial carcinoma, a squamous cell carcinoma, an adenocarcinoma, abronchogenic carcinoma, a renal cell carcinoma, a hepatoma, a bile ductcarcinoma, a neuroendocrine cancer, a carcinoid tumor, diffuse typegiant cell tumor, and glioblastoma.

In one embodiment, the cancer is a hematological cancer.

In one embodiment, the inflammatory disease is arthritis.

In one embodiment, the subject is a human. In one embodiment, thesubject is identified as having or being at risk of having a PI3K-gammamediated disorder via the use of a biomarker.

In one embodiment, the therapeutically effective dose is about 2 mg,about 1-3 mg, about 1-5 mg, about 1-10 mg, about 0.5-20 mg, about 0.1-50mg per day, about 0.1-75 mg per day, about 0.1-100 mg per day, about0.1-250 mg per day, about 0.1-500 mg per day, about 0.1-1000 mg per day,about 1-50 mg per day, about 1-75 mg per day, about 1-100 mg per day,about 1-250 mg per day, about 1-500 mg per day, about 1-1000 mg per day,about 10-50 mg per day, about 10-75 mg per day, about 10-100 mg per day,about 10-250 mg per day, about 10-500 mg per day, about 10-1000 mg perday, about 100-500 mg per day, or about 100-1000 mg per day. In oneembodiment, the therapeutically effective dose is about 0.029 mg/kg,about 0.014-0.14 mg/kg, about 0.02-0.04 mg/kg, about 0.01-0.05 mg/kg,about 0.01-0.1, or about 0.01-0.5 mg/kg. In one embodiment, the compoundis administered once every two days. In one embodiment, wherein thecompound is administered once per day. In one embodiment, the compoundis administered twice per day.

In one embodiment, the compound is administered at a dose such that thelevel of the compound in the subject is higher than the compound's IC50of PI3K-gamma inhibition during at least 70%, 80%, 90%, 95%, 97%, 98%,or 99% of a selected time period, e.g., 6 hours, 12 hours, 24 hours, or48 hours immediately following the administration. In one embodiment,the compound is administered at a dose such that the level of thecompound in the subject is higher than the compound's IC90 of PI3K-gammainhibition during at least 50%, 60%, 70%, 80%, 90%, 95%, 97%, 98%, or99% of a selected time period, e.g., 6 hours, 12 hours, 24 hours, or 48hours, immediately following the administration. In one embodiment, thecompound is administered at a dose such that the level of the compoundin the subject does not rise higher than the compound's IC20 or IC50 ofPI3K-delta inhibition within a selected time period, e.g., 6 hours, 12hours, 24 hours, or 48 hours, immediately following the administration.In one embodiment, the level of the compound is measured from thesubject's plasma. In one embodiment, the level of the compound ismeasured from the subject's tissue. In one embodiment, the compound isadministered at a dose such that it provides at least 50% inhibition ofPI3K-gamma in the subject but less than 10% or 20% inhibition ofPI3K-delta in the subject.

In one embodiment, the subject is a human and the compound has a halflife of about 10-13 hours in the subject. In one embodiment, the methodfurther comprises administering to the subject a second therapeuticagent that is a P-gp substrate. In one embodiment, the secondtherapeutic agent is Norvir (ritonavir).

PI3K-δ and/or PI3K-γ Activities

PI3K-δ has roles in impairments of B-cell signaling and development,antibody production, T-cell function, Th1 and Th2 differentiation, andmast and basophil degranulation. Without being limited by a particulartheory, PI3K-γ has roles in T-cell function, neutrophil and macrophagerecruitment, macrophage activation, neutrophil oxidative burst, anddendritic cell migration. Inhibition of PI3K-δ and/or PI3K-γ isoformscan result in efficacy against inflammation and cancer, e.g., inarthritis, asthma, multiple sclerosis (MS), and tumor models. Forexample, deficiency in PI3K-δ and/or PI3K-γ can result in efficacy inK/B×N arthritis model (Kyburz et al., Springer Semin. Immunopathology,2003, 25, 79-90) or K/B×N serum transfer model of arthritis (Randis etal., Eur. J. Immunol., 2008, 38(5), 1215-24), where it is shown thatrecognition of the immune complexes depends on both PI3K-δ and PI3K-γ,whereas cell migration is dependent on PI3K-γ. Deficiency in PI3K-δ orPI3K-γ can also result in efficacy in murine ovalbumin (OVA) inducedallergic asthma model (Lee et al., FASEB 1, 2006, 20, 455-65; Takeda etal., J. Allergy Clin. Immunol., 2009; 123, 805-12), where it is shownthat inhibition of either PI3K-δ or PI3K-γ inhibits ovalbumin inducedlung infiltration and improves airway responsiveness. Deficiency inPI3K-δ or PI3K-γ can also result in efficacy in murine experimentalautoimmune encephalomyelitis (model for MS), where it is shown thatPI3K-γ deletion may provide better efficacy as compared to PI3K-δdeletion (Haylock-Jacob et al., J. Autoimmunity, 2011, 36, 278-87;Comerford et al., PLOS One, 2012, 7, e45095), including reduction inT-cell receptor induced CD4⁺ T cell activation, leukocyte infiltrationand Th1/Th17 responses, and dendritic cell migration (Comerfold, PLOSOne, 2012, 7, e45095). Furthermore, inhibition of PI3K-γ can also resultin decreased tumor inflammation and growth (e.g., Lewis lung carcinomamodel, Schmid et al., Cancer Cell, 2011, 19(6), 715-27). PI3K-γ deletioncombined with PI3K-δ deletion results in increased survival in T-cellacute lymphoblastic leukemia (T-ALL) (Subramaniam et al., Cancer Cell,2012, 21, 459-72). Inhibitors of both PI3K-δ and PI3K-γ are also shownto be efficacious in PTEN-deleted T-ALL cell line (MOLT-4). In theabsence of PTEN phosphatase tumor suppressor function, PI3K-δ or PI3K-γalone can support the development of leukemia, whereas inactivation ofboth isoforms suppresses tumor formation. Thus, inhibitors of PI3K-δand/or PI3K-γ can be useful in treating inflammation, such as arthritis,allergic asthma, and MS; and in treating cancer, for example, due toeffects such as reductions in solid tumor associated inflammation,angiogenesis and tumor progression.

The importance of PI3K-δ in the development and function of B-cells issupported from inhibitor studies and genetic models. PI3K-δ is animportant mediator of B-cell receptor (BCR) signaling, and is upstreamof AKT, calcium flux, PLCγ, MAP kinase, P70S6k, and FOXO3a activation.PI3K-δ is also important in IL4R, SIP, and CXCR5 signaling, and has beenshown to modulate responses to toll-like receptors 4 and 9. Inhibitorsof PI3K-δ have shown the importance of PI3K-δ in B-cell development(Marginal zone and B1 cells), B-cell activation, chemotaxis, migrationand homing to lymphoid tissue, and in the control of immunoglobulinclass switching leading to the production of IgE. Clayton E et al.(2002) J Exp Med. 196(6):753-63; Bilancio A, et al. (2006) Blood107(2):642-50; Okkenhaug K. et al. (2002) Science 297(5583):1031-4;Al-Alwan M M et al. (2007) J Immunol. 178(4):2328-35; Zhang T T, et al.(2008) J Allergy Clin Immunol. 2008; 122(4):811-819.e2; Srinivasan L, etal. (2009) Cell 139(3):573-86).

In T-cells, PI3K-δ has been demonstrated to have a role in T-cellreceptor and cytokine signaling, and is upstream of AKT, PLCγ, andGSK3b. In PI3K-δ deletion or kinase-dead knock-in mice, or in inhibitorstudies, T-cell defects including proliferation, activation, anddifferentiation have been observed, leading to reduced T helper cell 2(TH2) response, memory T-cell specific defects (DTH reduction), defectsin antigen dependent cellular trafficking, and defects inchemotaxis/migration to chemokines (e.g., SIP, CCR7, CD62L). (Garçon F.et al. (2008) Blood 111(3):1464-71; Okkenhaug K et al. (2006). JImmunol. 177(8):5122-8; Soond D R, et al. (2010) Blood 115(11):2203-13;Reif K, (2004). J Immunol. 2004; 173(4):2236-40; Ji H. et al. (2007)Blood 110(8):2940-7; Webb L M, et al. (2005) J Immunol. 175(5):2783-7;Liu D, et al. (2010) J Immunol. 184(6):3098-105; Haylock-Jacobs S, etal. (2011) J Autoimmun. 2011; 36(3-4):278-87; Jarmin S J, et al. (2008)J Clin Invest. 118(3):1154-64).

Numerous publications support roles of PI3K-δ and PI3K-γ in thedifferentiation, maintenance, and activation of immune and malignantcells, as described in more detail herein.

PI3K-δ and PI3K-γ isoforms are preferentially expressed in leukocyteswhere they have distinct and non-overlapping roles in immune celldevelopment and function. See, e.g., PURI and GOLD, “Selectiveinhibitors of phosphoinositide 3-kinase delta: modulators of B-cellfunction with potential for treating autoimmune inflammatory disease andB-cell malignancies,” Front. Immunol. 3:256 (2012); BUITENHUIS et al.,“The role of the PI3K-PKB signaling module in regulation ofhematopoiesis,” Cell Cycle 8(4):560-566 (2009); HOELLENRIEGEL andBURGER, “Phosphoinositide 3′-kinase delta: turning off BCR signaling inChronic Lymphocytic Leukemia,” Oncotarget 2(10):737-738 (2011); HIRSCHet al., “Central Role for G Protein-Coupled Phosphoinositide 3-Kinase γin Inflammation,” Science 287:1049-1053 (2000); L I et al., “Roles ofPLC-β2 and -β3 and PI3Kγ in Chemoattractant-Mediated SignalTransduction,” Science 287:1046-1049 (2000); SASAKI et al., “Function ofPI3Kγ in Thymocyte Development, T Cell Activation, and NeutrophilMigration,” Science 287:1040-1046 (2000); CUSHING et al., “PI3Kδ andPI3Kγ as Targets for Autoimmune and Inflammatory Diseases,” J. Med.Chem. 55:8559-8581 (2012); MAXWELL et al., “Attenuation ofphosphoinositide 3-kinase δ signaling restrains autoimmune disease,” J.Autoimmun. 38:381-391 (2012); HAYLOCK-JACOBS et al., “PI3Kδ drives thepathogenesis of experimental autoimmune encephalomyelitis by inhibitingeffector T cell apoptosis and promoting Th17 differentiation,” J.Autoimmun. 36:278-287 (2011); SOOND et al., “PI3K p110δ regulates T-cellcytokine production during primary and secondary immune responses inmice and humans,” Blood 115(11):2203-2213 (2010); ROLLER et al.,“Blockade of Phosphatidylinositol 3-Kinase (PI3K)δ or PI3Kγ ReducesIL-17 and Ameliorates Imiquimod-Induced Psoriasis-like Dermatitis,” J.Immunol. 189:4612-4620 (2012); CAMPS et al., “Blockade of PI3Kγsuppresses joint inflammation and damage in mouse models of rheumatoidarthritis,” Nat. Med. 11(9):936-943 (2005). As key enzymes in leukocytesignaling, PI3K-δ and PI3K-γ facilitate normal B-cell, T-cell andmyeloid cell functions including differentiation, activation, andmigration. See, e.g., HOELLENRIEGEL and BURGER, “Phosphoinositide3′-kinase delta: turning off BCR signaling in Chronic LymphocyticLeukemia,” Oncotarget 2(10):737-738 (2011); CUSHING et al., “PI3Kδ andPI3Kγ as Targets for Autoimmune and Inflammatory Diseases,” J. Med.Chem. 55:8559-8581 (2012). PI3K-δ or PI3K-γ activity is critical forpreclinical models of autoimmune and inflammatory diseases. See, e.g.,HIRSCH et al., “Central Role for G Protein-Coupled Phosphoinositide3-Kinase γ in Inflammation,” Science 287:1049-1053 (2000); L I et al.,“Roles of PLC-β2 and -β3 and PI3Kγ in Chemoattractant-Mediated SignalTransduction,” Science 287:1046-1049 (2000); SASAKI et al., “Function ofPI3Kγ in Thymocyte Development, T Cell Activation, and NeutrophilMigration,” Science 287:1040-1046 (2000); CUSHING et al., “PI3Kδ andPI3Kγ as Targets for Autoimmune and Inflammatory Diseases,” J. Med.Chem. 55:8559-8581 (2012); MAXWELL et al., “Attenuation ofphosphoinositide 3-kinase δ signaling restrains autoimmune disease,” J.Autoimmun. 38:381-391 (2012); HAYLOCK-JACOBS et al., “PI3Kδ drives thepathogenesis of experimental autoimmune encephalomyelitis by inhibitingeffector T cell apoptosis and promoting Th17 differentiation,” J.Autoimmun. 36:278-287 (2011); SOOND et al., “PI3K p110δ regulates T-cellcytokine production during primary and secondary immune responses inmice and humans,” Blood 115(11):2203-2213 (2010); ROLLER et al.,“Blockade of Phosphatidylinositol 3-Kinase (PI3K)δ or PI3Kγ ReducesIL-17 and Ameliorates Imiquimod-Induced Psoriasis-like Dermatitis,” J.Immunol. 189:4612-4620 (2012); CAMPS et al., “Blockade of PI3Kγsuppresses joint inflammation and damage in mouse models of rheumatoidarthritis,” Nat. Med. 11(9):936-943 (2005). Given the key role forPI3K-δ and PI3K-γ in immune function, inhibitors of the PI3K-δ and/or γhave therapeutic potential in immune-related inflammatory or neoplasticdiseases.

PI3K-δ and PI3K-γ are central to the growth and survival of B- andT-cell malignancies and inhibition of these isoforms may effectivelylimit these diseases. See, e.g., SUBRAMANIAM et al., “TargetingNonclassical Oncogenes for Therapy in T-ALL,” Cancer Cell 21:459-472(2012); LANNUTTI et al., “CAL-101 a p110δ selectivephosphatidylinositol-3-kinase inhibitor for the treatment of B-cellmalignancies, inhibits PI3K signaling and cellular viability,” Blood117(2):591-594 (2011). PI3K-δ and PI3K-γ support the growth and survivalof certain B-cell malignancies by mediating intracellular BCR signalingand interactions between the tumor cells and their microenvironment.See, e.g., PURI and GOLD, “Selective inhibitors of phosphoinositide3-kinase delta: modulators of B-cell function with potential fortreating autoimmune inflammatory disease and B-cell malignancies,”Front. Immunol. 3:256 (2012); HOELLENRIEGEL et al., “Thephosphoinositide 3′-kinase delta inhibitor, CAL-101, inhibits B-cellreceptor signaling and chemokine networks in chronic lymphocyticleuckemia,” Blood 118(13): 3603-3612 (2011); BURGER, “Inhibiting B-CellReceptor Signaling Pathways in Chronic Lymphocytic Leukemia,” Curr.Mematol. Malig. Rep. 7:26-33 (2012). Increased BCR signaling is acentral pathologic mechanism of B-cell malignancies and PI3K activationis a direct consequence of BCR pathway activation. See, e.g., BURGER,“Inhibiting B-Cell Receptor Signaling Pathways in Chronic LymphocyticLeukemia,” Curr. Mematol. Malig. Rep. 7:26-33 (2012); HERISHANU et al.,“The lymph node microenvironment promotes B-cell receptor signaling,NF-κB activation, and tumor proliferation in chronic lymphocyticleukemia,” Blood 117(2):563-574 (2011); DAVIS et al., “Chronic activeB-cell-receptor signaling in diffuse large B-cell lymphoma,” Nature463:88-92 (2010); PIGHI et al., “Phospho-proteomic analysis of mantlecell lymphoma cells suggests a pro-survival role of B-cell receptorsignaling,” Cell Oncol. (Dordr) 34(2):141-153 (2011); RIZZATTI et al.,“Gene expression profiling of mantle cell lymphoma cells revealsaberrant expression of genes from the PI3K-AKT, WNT and TGFβ signalingpathways,” Brit. J. Haematol. 130:516-526 (2005); MARTINEZ et al., “TheMolecular Signature of Mantle Cell Lymphoma Reveals Multiple SignalsFavoring Cell Survival,” Cancer Res. 63:8226-8232 (2003). Interactionsbetween malignant B-cells and supporting cells (eg, stromal cells,nurse-like cells) in the tumor microenvironment are important for tumorcell survival, proliferation, homing, and tissue retention. See, e.g.,BURGER, “Inhibiting B-Cell Receptor Signaling Pathways in ChronicLymphocytic Leukemia,” Curr. Mematol. Malig. Rep. 7:26-33 (2012);HERISHANU et al., “The lymph node microenvironment promotes B-cellreceptor signaling, NF-κB activation, and tumor proliferation in chroniclymphocytic leukemia,” Blood 117(2):563-574 (2011); KURTOVA et al.,“Diverse marrow stromal cells protect CLL cells from spontaneous anddrug-induced apoptosis: development of a reliable and reproduciblesystem to assess stromal cell adhesion-mediated drug resistance,” Blood114(20): 4441-4450 (2009); BURGER et al., “High-level expression of theT-cell chemokines CCL3 and CCL4 by chronic lymphocytic leukemia B cellsin nurselike cell cocultures and after BCR stimulation,” Blood 113(13)3050-3058 (2009); QUIROGA et al., “B-cell antigen receptor signalingenhances chronic lymphocytic leukemia cell migration and survival:specific targeting with a novel spleen tyrosine kinase inhibitor, R406,”Blood 114(5):1029-1037 (2009). Inhibiting PI3K-δ,γ with an inhibitor incertain malignant B-cells can block the BCR-mediated intracellularsurvival signaling as well as key interactions with theirmicroenvironment that are critical for their growth.

PI3K-δ and PI3K-γ also play a direct role in the survival andproliferation of certain T-cell malignancies. See, e.g., SUBRAMANIAM etal., “Targeting Nonclassical Oncogenes for Therapy in T-ALL,” CancerCell 21:459-472 (2012). Aberrant PI3K-δ and PI3K-γ activity provides thesignals necessary for the development and growth of certain T-cellmalignancies. While BTK is expressed in B-cells, it is not expressed inT-cells, and therefore BTK is not a viable target for the treatment ofT-cell malignancies. See, e.g., NISITANI et al., “Posttranscriptionalregulation of Bruton's tyrosine kinase expression in antigenreceptor-stimulated splenic B cells,” PNAS 97(6):2737-2742 (2000); DEWEERS et al., “The Bruton's tyrosine kinase gene is expressed throughoutB cell differentiation, from early precursor B cell stages precedingimmunoglobulin gene rearrangement up to mature B cell stages,” Eur. J.Immunol. 23:3109-3114 (1993); SMITH et al., “Expression of Bruton'sAgammaglobulinemia Tyrosine Kinase Gene, BTK, Is SelectivelyDown-Regulated in T Lymphocytes and Plasma Cells,” J. Immunol.152:557-565 (1994). PI3K-δ and/or γ inhibitors may have uniquetherapeutic potential in T-cell malignancies.

In neutrophils, PI3K-δ, along with PI3K-γ, contribute to the responsesto immune complexes, FCγRII signaling, including migration andneutrophil respiratory burst. Human neutrophils undergo rapid inductionof PIP3 in response to formyl peptide receptor (FMLP) or complementcomponent C5a (C5a) in a PI3K-γ dependent manner, followed by a longerPIP3 production period that is PI3K-δ dependent, and is essential forrespiratory burst. The response to immune complexes is contributed byPI3K-δ, PI3K-γ, and PI3K-β, and is an important mediator of tissuedamage in models of autoimmune disease (Randis T M et al. (2008) Eur JImmunol. 38(5):1215-24; Pinho V, (2007) J Immunol. 179(11):7891-8; SadhuC. et al. (2003) J Immunol. 170(5):2647-54; Condliffe A M et al. (2005)Blood 106(4):1432-40). It has been reported that in certain autoimmunediseases, preferential activation of PI3K-β may be involved (Kulkarni etal., Immunology (2011) 4(168) ra23: 1-11). It was also reported thatPI3K-β-deficient mice were highly protected in an FcγR-dependent modelof autoantibody-induced skin blistering and partially protected in anFcγR-dependent model of inflammatory arthritis, whereas combineddeficiency of PI3K-β and PI3K-δ resulted in near complete protection ininflammatory arthritis (Id.).

In macrophages collected from patients with chronic obstructivepulmonary disease (COPD), glucocorticoid responsiveness can be restoredby treatment of the cells with inhibitors of PI3K-δ. Macrophages alsorely on PI3K-δ and PI3K-γ for responses to immune complexes through thearthus reaction (FCγR and C5a signaling) (Randis T M, et al. (2008) EurJ Immunol. 38(5):1215-24; Marwick J A et al. (2009) Am J Respir CritCare Med. 179(7):542-8; Konrad S, et al. (2008) J Biol Chem.283(48):33296-303).

Theophylline increases histone deacetylase-2 and corticosteroidsensitivity in vitro and in smoking mice in vivo by inhibiting PI3kinase (e.g., PI3K-delta). PI3K is activated in COPD lungs and certainPI3K inhibitors have been shown to mimic the effects of theophylline inreversing corticosteroid resistance. Yasuo, T., et al., Am J Respir CritCare Med 2010; 182:897-904. While not wishing to be bound by theory, arationale for the use of PI3K inhibitors (e.g., compounds providedherein) to treat COPD is that a PI3K inhibitor can increase thecorticosteroid sensitivity in a subject.

In mast cells, stem cell factor-(SCF) and IL3-dependent proliferation,differentiation and function are PI3K-δ dependent, as is chemotaxis. Theallergen/IgE crosslinking of FCγR1 resulting in cytokine release anddegranulation of the mast cells is severely inhibited by treatment withPI3K-δ inhibitors, suggesting a role for PI3K-δ in allergic disease (AliK et al. (2004) Nature 431(7011):1007-11; Lee K S, et al. (2006) FASEBJ. 20(3):455-65; Kim M S, et al. (2008) Trends Immunol. 29(10):493-501).

Natural killer (NK) cells are dependent on both PI3K-δ and PI3K-γ forefficient migration towards chemokines including CXCL10, CCL3, S1P andCXCL12, or in response to LPS in the peritoneum (Guo H, et al. (2008) JExp Med. 205(10):2419-35; Tassi I, et al. (2007) Immunity 27(2):214-27;Saudemont A, (2009) Proc Natl Acad Sci USA. 106(14):5795-800; Kim N, etal. (2007) Blood 110(9): 3202-8).

The roles of PI3K-δ and PI3K-γ in the differentiation, maintenance, andactivation of immune cells support a role for these enzymes ininflammatory disorders ranging from autoimmune diseases (e.g.,rheumatoid arthritis, multiple sclerosis) to allergic inflammatorydisorders, such as asthma, and inflammatory respiratory disease, such asCOPD. Extensive evidence is available in experimental animal models, orcan be evaluated using art-recognized animal models. In an embodiment,described herein is a method of treating inflammatory disorders rangingfrom autoimmune diseases (e.g., rheumatoid arthritis, multiplesclerosis) to allergic inflammatory disorders, such as asthma and COPDusing a compound described herein.

For example, inhibitors of PI3K-δ and/or -γ have been shown to haveanti-inflammatory activity in several autoimmune animal models forrheumatoid arthritis (Williams, O. et al. (2010) Chem Biol,17(2):123-34; WO 2009/088986; WO2009/088880; WO 2011/008302; eachincorporated herein by reference). PI3K-δ is expressed in the RAsynovial tissue (especially in the synovial lining which containsfibroblast-like synoviocytes (FLS), and selective PI3K-δ inhibitors havebeen shown to be effective in inhibiting synoviocyte growth and survival(Bartok et al. (2010) Arthritis Rheum 62 Suppl 10:362). Several PI3K-δand -γ inhibitors have been shown to ameliorate arthritic symptoms(e.g., swelling of joints, reduction of serum-induced collagen levels,reduction of joint pathology and/or inflammation), in art-recognizedmodels for RA, such as collagen-induced arthritis and adjuvant inducedarthritis (WO 2009/088986; WO2009/088880; WO 2011/008302; eachincorporated herein by reference).

The role of PI3K-δ has also been shown in models of T-cell dependentresponse, including the DTH model. In the murine experimental autoimmuneencephalomyelitis (EAE) model of multiple sclerosis, the PI3K-γ/δ-doublemutant mice are resistant. PI3K-δ inhibitors have also been shown toblock EAE disease induction and development of TH-17 cells both in vitroand in vivo (Haylock-Jacobs, S. et al. (2011) J. Autoimmunity36(3-4):278-87).

Systemic lupus erythematosus (SLE) is a complex disease that atdifferent stages requires memory T-cells, B-cell polyclonal expansionand differentiation into plasma cells, and the innate immune response toendogenous damage associated molecular pattern molecules (DAMPS), andthe inflammatory responses to immune complexes through the complementsystem as well as the Fc receptors. The role of PI3K-δ and PI3K-γtogether in these pathways and cell types suggest that blockade with aninhibitor would be effective in these diseases. A role for PI3K in lupusis also predicted by two genetic models of lupus. The deletion ofphosphatase and tensin homolog (PTEN) leads to a lupus-like phenotype,as does a transgenic activation of Class 1A PI3Ks, which includesPI3K-δ. The deletion of PI3K-γ in the transgenically activated class 1Alupus model is protective, and treatment with a PI3K-γ selectiveinhibitor in the murine MLR/lpr model of lupus improves symptoms(Barber, D F et al. (2006) J. Immunol. 176(1): 589-93).

In allergic disease, PI3K-δ has been shown by genetic models and byinhibitor treatment to be essential for mast-cell activation in apassive cutaneous anaphalaxis assay (Ali K et al. (2008) J Immunol.180(4):2538-44; Ali K, (2004) Nature 431(7011):1007-11). In a pulmonarymeasure of response to immune complexes (Arthus reaction) a PI3K-δknockout is resistant, showing a defect in macrophage activation and C5aproduction. Knockout studies and studies with inhibitors for both PI3K-δand PI3K-γ support a role for both of these enzymes in the ovalbumininduced allergic airway inflammation and hyper-responsiveness model (LeeK S et al. (2006) FASEB J. 20(3):455-65). Reductions of infiltration ofeosinophils, neutrophils, and lymphocytes as well as TH2 cytokines (IL4,IL5, and IL13) were seen with both PI3K-δ specific and dual PI3K-δ andPI3K-γ inhibitors in the Ova induced asthma model (Lee K S et al. (2006)J Allergy Clin Immunol 118(2):403-9).

PI3K-δ and PI3K-γ inhibition can be used in treating COPD. In the smokedmouse model of COPD, the PI3K-δ knockout does not develop smoke inducedglucocorticoid resistance, while wild-type and PI3K-γ knockout mice do.An inhaled formulation of dual PI3K-δ and PI3K-γ inhibitor blockedinflammation in a LPS or smoke COPD models as measured by neutrophiliaand glucocorticoid resistance (Doukas J, et al. (2009) J Pharmacol ExpTher. 328(3):758-65).

PI3K-δ and/or PI3K-γ Isoforms in Certain Cancers

Class I PI3Ks, particularly PI3K-δ and PI3K-γ isoforms, are alsoassociated with cancers (reviewed, e.g., in Vogt, P K et al. (2010) CurrTop Microbiol Immunol. 347:79-104; Fresno Vara, J A et al. (2004) CancerTreat Rev. 30(2):193-204; Zhao, L and Vogt, P K. (2008) Oncogene27(41):5486-96). Inhibitors of PI3K, e.g., PI3K-δ and/or PI3K-γ, havebeen shown to have anti-cancer activity (e.g., Courtney, K D et al.(2010) J Clin Oncol. 28(6):1075-1083); Markman, B et al. (2010) AnnOncol. 21(4):683-91; Kong, D and Yamori, T (2009) Curr Med Chem.16(22):2839-54; Jimeno, A et al. (2009) J Clin Oncol. 27:156s (suppl;abstr 3542); Flinn, I W et al. (2009) J Clin Oncol. 27:156s (suppl;abstr 3543); Shapiro, G et al. (2009) J Clin Oncol. 27:146s (suppl;abstr 3500); Wagner, A J et al. (2009) J Clin Oncol. 27:146s (suppl;abstr 3501); Vogt, P K et al. (2006) Virology 344(1):131-8; Ward, S etal. (2003) Chem Biol. 10(3):207-13; WO 2011/041399; US 2010/0029693; US2010/0305096; US 2010/0305084; each incorporated herein by reference).

In one embodiment, described herein is a method of treating cancer. Inone embodiment, provided herein is a method of treating a hematologicalcancer comprising administering a pharmaceutically effective amount of acompound provided herein to a subject in need thereof. In oneembodiment, provided herein is a method of treating a solid tumorcomprising administering a pharmaceutically effective amount of acompound provided herein to a subject in need thereof. Types of cancerthat can be treated with an inhibitor of PI3K (particularly, PI3K-δand/or PI3K-γ) include, e.g., leukemia, chronic lymphocytic leukemia,acute myeloid leukemia, chronic myeloid leukemia (e.g., Salmena, L etal. (2008) Cell 133:403-414; Chapuis, N et al. (2010) Clin Cancer Res.16(22):5424-35; Khwaja, A (2010) Curr Top Microbiol Immunol.347:169-88); lymphoma, e.g., non-Hodgkin's lymphoma (e.g., Salmena, L etal. (2008) Cell 133:403-414); lung cancer, e.g., non-small cell lungcancer, small cell lung cancer (e.g., Herrera, V A et al. (2011)Anticancer Res. 31(3):849-54); melanoma (e.g., Haluska, F et al. (2007)Semin Oncol. 34(6):546-54); prostate cancer (e.g., Sarker, D et al.(2009) Clin Cancer Res. 15(15):4799-805); glioblastoma (e.g., Chen, J Set al. (2008) Mol Cancer Ther. 7:841-850); endometrial cancer (e.g.,Bansal, N et al. (2009) Cancer Control. 16(1):8-13); pancreatic cancer(e.g., Furukawa, T (2008) J Gastroenterol. 43(12):905-11); renal cellcarcinoma (e.g., Porta, C and Figlin, R A (2009) J Urol.182(6):2569-77); colorectal cancer (e.g., Saif, M W and Chu, E (2010)Cancer J. 16(3):196-201); breast cancer (e.g., Torbett, N E et al.(2008) Biochem J. 415:97-100); thyroid cancer (e.g., Brzezianska, E andPastuszak-Lewandoska, D (2011) Front Biosci. 16:422-39); and ovariancancer (e.g., Mazzoletti, M and Broggini, M (2010) Curr Med Chem.17(36):4433-47).

Numerous publications support a role of PI3K-δ and PI3K-γ in treatinghematological cancers. PI3K-δ and PI3K-γ are highly expressed in theheme compartment, and solid tumors, including prostate, breast andglioblastomas (Chen J. S. et al. (2008) Mol Cancer Ther. 7(4):841-50;Ikeda H. et al. (2010) Blood 116(9): 1460-8).

In hematological cancers including acute myeloid leukemia (AML),multiple myeloma (MM), and chronic lymphocytic leukemia (CLL),overexpression and constitutive activation of PI3K-δ supports the modelthat PI3K-δ inhibition would be therapeutic Billottet C, et al. (2006)Oncogene 25(50):6648-59; Billottet C, et al. (2009) Cancer Res.69(3):1027-36; Meadows, S A, 52^(nd) Annual ASH Meeting and Exposition;2010 Dec. 4-7; Orlando, Fla.; Ikeda H, et al. (2010) Blood116(9):1460-8; Herman S E et al. (2010) Blood 116(12):2078-88; Herman SE et al. (2011). Blood 117(16):4323-7.

In one embodiment, described herein is a method of treatinghematological cancers including, but not limited to acute myeloidleukemia (AML), multiple myeloma (MM), and chronic lymphocytic leukemia(CLL).

A PI3K-δ inhibitor (CAL-101) has been evaluated in a phase 1 trial inpatients with haematological malignancies, and showed activity in CLL inpatients with poor prognostic characteristics. In CLL, inhibition ofPI3K-δ not only affects tumor cells directly, but it also affects theability of the tumor cells to interact with their microenvironment. Thismicroenvironment includes contact with and factors from stromal cells,T-cells, nurse like cells, as well as other tumor cells. CAL-101suppresses the expression of stromal and T-cell derived factorsincluding CCL3, CCL4, and CXCL13, as well as the CLL tumor cells'ability to respond to these factors. CAL-101 treatment in CLL patientsinduces rapid lymph node reduction and redistribution of lymphocytesinto the circulation, and affects tonic survival signals through theBCR, leading to reduced cell viability, and an increase in apoptosis.Single agent CAL-101 treatment was also active in mantle cell lymphomaand refractory non Hodgkin's lymphoma (Furman, R R, et al. 52^(nd)Annual ASH Meeting and Exposition; 2010 Dec. 4-7; Orlando, Fla.;Hoellenriegel, J, et al. 52^(nd) Annual ASH Meeting and Exposition; 2010Dec. 4-7; Orlando, Fla.; Webb, H K, et al. 52^(nd) Annual ASH Meetingand Exposition; 2010 Dec. 4-7; Orlando, Fla.; Meadows, et al. 52^(11d)Annual ASH Meeting and Exposition; 2010 Dec. 4-7; Orlando, Fla.; Kahl,B, et al. 52^(nd) Annual ASH Meeting and Exposition; 2010 Dec. 4-7;Orlando, Fla.; Lannutti B J, et al. (2011) Blood 117(2):591-4).

PI3K-δ inhibitors have shown activity against PI3K-δ positive gliomas invitro (Kashishian A, et al. Poster presented at: The AmericanAssociation of Cancer Research 102^(nd) Annual Meeting; 2011 Apr. 2-6;Orlando, Fla.). In this subset of tumors, treatment with the PI3K-δinhibitor either alone or in combination with a cytotoxic agent can beeffective.

Another mechanism for PI3K-δ inhibitors to have an effect in solidtumors involves the tumor cells' interaction with theirmicro-environment. PI3K-δ, PI3K-γ, and PI3K-β are expressed in theimmune cells that infiltrate tumors, including tumor infiltratinglymphocytes, macrophages, and neutrophils. PI3K-δ inhibitors can modifythe function of these tumor-associated immune cells and how they respondto signals from the stroma, the tumor, and each other, and in this wayaffect tumor cells and metastasis (Hoellenriegel, J, et al. 52^(nd)Annual ASH Meeting and Exposition; 2010 Dec. 4-7; Orlando, Fla.).

PI3K-δ is also expressed in endothelial cells. It has been shown thattumors in mice treated with PI3K-δ selective inhibitors are killed morereadily by radiation therapy. In this same study, capillary networkformation is impaired by the PI3K inhibitor, and it is postulated thatthis defect contributes to the greater killing with radiation. PI3K-δinhibitors can affect the way in which tumors interact with theirmicroenvironment, including stromal cells, immune cells, and endothelialcells and be therapeutic either on its own or in conjunction withanother therapy (Meadows, S A, et al. Paper presented at: 52^(nd) AnnualASH Meeting and Exposition; 2010 Dec. 4-7; Orlando, Fla.; Geng L, et al.(2004) Cancer Res. 64(14):4893-9).

Accordingly, provided herein is a method of treating or preventing acancer or disease (including but not limited to a hematologicmalignancy, or a specific type or sub-type of cancer or disease, such asa specific type or sub-type of hematologic malignancy), with a PI3K-γselective inhibitor, wherein the adverse effects associated withadministration of inhibitors for other isoform(s) of PI3K (e.g., PI3K-αand/or PI3K-β) are reduced. In one embodiment, provided herein is amethod of treating or preventing the cancer or disease, with a PI3K-γselective inhibitor, at a lower (e.g., by about 10%, by about 20%, byabout 30%, by about 40%, by about 50%, by about 60%, by about 70%, or byabout 80%) dose as compared to treatment with a PI3K-γ non-selective orless selective PI3K-γ inhibitor (e.g., a PI3Kpan inhibitors, e.g.,inhibiting PI3K-α, β, δ, and γ).

The role of PI3K-γ pathway in promoting myeloid cell trafficking totumors and the role of blockade of p100γ in suppression of tumorinflammation and growth in breast cancer, pancreatic cancer, and lungcancer are reported, for example, in Schmid et al. (2011) Cancer Cell19, 715-727, the entirety of which is incorporated herein by reference.In one embodiment, provided herein is a method of treating or preventingpancreatic cancer with a PI3K inhibitor.

In another embodiment, provided herein is a method of treating orpreventing breast cancer with a PI3K inhibitor. In yet anotherembodiment, provided herein is a method of treating or preventing lungcancer with a PI3K inhibitor. In one embodiment, the PI3K inhibitor is aPI3K-γ inhibitor, selective or non-selective over one or more other PI3Kisoform(s). In one embodiment, the PI3K inhibitor is a PI3K-γ selectiveinhibitor.

While not wishing to be bound by theory, it is believed that tumorgrowth is influenced by two classes of immune cells in the tumormicroenvironment: effector cells which include cytotoxic cells and M1macrophages, and which have anti-tumor activity, and suppressor cells,which include M2 macrophages, MDSC (myeloid derived suppressor cell),Tregs (regulatory T cell), and regulatory dendritic cells, which havepro-tumor activity because they inhibit the effector cells. An abundanceof suppressor cells can lead to tumor immune tolerance, and enhancementof tumor growth.

Certain of these cell types are briefly described. M1 denotes apro-inflammatory (anti-tumor) phenotype of a MDSC or TAM. M2 denotes ananti-inflammatory (pro-tumor) phenotype of a MDSC or TAM.

PI3K-γ is not expressed in at least some cancer cell types. Schmid etal., 2011, Cancer Cell 19. Accordingly, in some embodiments, the PI3K-γinhibitor reduces cancer cell growth without having a substantial directeffect on the cancer cell itself. For instance, in some embodiments, thePI3K-γ inhibitor inhibits cancer cell growth through changes in thetumor microenvironment, e.g., the immune cells in close proximity to thecancer cells.

Evidence provided in the Examples herein, combined with evidence in theliterature, support the idea that a PI3K-γ inhibitor can reduce tumorassociated myeloid cells. For instance, in PI3K-γ-deficient mice,tumor-associated myeloid cells are reduced. Schmid et al., 2011, CancerCell 19. Together, these data indicate that a large class of PI3K-γinhibitors should reduce tumor associated myeloid cells, therebyincreasing the immune response against cancer cells, and treating thecancer. While not wishing to be bound by theory, a PI3K-γ may operatethrough the following mechanism. PI3K-γ signaling may tilt the balanceof immune cells towards pro-tumor M2 cells and away from anti-tumor M1cells, by inducing expression of immunosuppressive, wound healing genessuch as Arginase1, TGFbeta1, PDGFBB, MMP9, and MMP13, and suppressingpro-inflammatory factors such as IL12, iNos, and interferon gamma.Blocking PI3K-γ signaling with an inhibitor tilts the balance towardsanti-tumor M1 cells by stimulating a T cell activating gene expressionprogram. Kaneda et al. PI3-kinase gamma controls the macrophage M1-M2switch, thereby promoting tumor immunosuppression and progression.[abstract]. In: Proceedings of the 105th Annual Meeting of the AmericanAssociation for Cancer Research; 2014 Apr. 5-9; San Diego, Calif.Philadelphia (Pa.): AACR; Cancer Res 2014; 74(19 Suppl):Abstract nr3650. doi:10.1158/1538-7445.AM2014-3650.

In some embodiments, a PI3K-γ inhibitor provided herein is administeredto a patient in order to block a homeostatic down-regulation of T cellresponse. While not wishing to be bound by theory, this may allow thebody to raise an effective immune response against the cancer cell.Exemplary agents of this type include immune checkpoint therapeutics,e.g., agents that act on CTLA-4, PD-1, or PD-L1, e.g., antibodies thatbind to CTLA-4, PD-1, or PD-L1. Immune checkpoint therapeutics aredescribed in more detail below.

In some embodiments, a PI3K-γ inhibitor provided herein is administeredto a patient in order to eliminate immunosuppressive cells in the tumormicroenvironment. The immunosuppressive cell may be, e.g., a Tregulatory cell (e.g., a cell that secretes mediators that induce CD8+cytotoxic T cell death); a Tumor-associated macrophage (TAM; e.g., anM2(pro-tumor) TAMS that blocks T cell activity and promotes angiogenesis);or a myeloid-derived suppressor cell (MDSC; e.g., a cell that secretesmediators that inhibit T cell differentiation and proliferation).

In some embodiments, a compound provided herein is administered to apatient in order to reduce the migration or differentiation of a tumorassociated myeloid cell. In some embodiments, the compound is a compoundthat shows single agent activity in a syngeneic model system. In someembodiments, the compound is administered in combination with a secondtherapeutic, as discussed herein. In some embodiments, theadministration results in a reduction in the level of MDSCs in the tumormicroenvironment; the level of M2 TAMS in the tumor microenvironment;the level of T-regulatory cells in the tumor microenvironment, or anycombination thereof. In some embodiments, the administration results inan unchanged or increased level of T-effector cells in the tumormicroenvironment. In embodiments, the administration results in anincrease in an immune response to the tumor, e.g., an increase in thelevels or tumor-attacking activity of cytotoxic T cells, M1 inflammatoryTAMs, or a combination thereof.

In some embodiments, an MDSC has one or more of the followingproperties: suppressing anti-tumor immune attack; inducingvascularization of the tumor; inducing ECM breakdown, e.g., which maycontribute to metastasis; and supporting tumor growth. Accordingly, insome embodiments, administration of a PI3K-γ inhibitor described hereininhibits one or more of these functions in an MDSC.

TAMs (tumor-associated macrophages) can also have one or more of thefollowing properties: suppressing anti-tumor immune attack; inducingvascularization of the tumor; inducing ECM breakdown, e.g., which maycontribute to metastasis; and supporting tumor growth. Accordingly, insome embodiments, administration of a PI3K-γ inhibitor as describedherein inhibits one or more of these functions in a TAM.

In embodiments, a PI3K-γ inhibitor is administered to a patient who hasreceived chemotherapy and/or radiation therapy. While not wishing to bebound by theory, in some embodiments, chemotherapy or radiation therapyresults in a wound healing response that leads to repopulation of thecancer site, e.g., tumor, with TAMs and MDSCs. Administering the PI3K-γinhibitor, in some embodiments, reduces the levels of TAMs and MDSCs inthe microenvironment, decreasing their support for tumor cell growthand/or allowing the immune system to attack the cancer cells. See ClaireE. Lewis, “Imaging immune cell infiltrating tumors in zebrafish”, AACRAnnual Meeting (Apr. 5, 2014).

While not wishing to be bound by theory, a rationale for the use of aPI3K gamma inhibitor as adjunct therapy to radiation is to prevent theaccumulation of tumor supporting myeloid cells into the radiated tumor,thus impairing tumor regrowth following radiation therapy. This issupported by work of Kioi et al. (2010) Clin Invest. 120(3):694-705showing that an inhibitor of myeloid cell migration into post-irradiatedtumors (e.g., AMD3100) blocked tumor vasculogenesis and tumor regrowth.

In certain embodiments, provided herein is a method of treating adisorder or disease provided herein, comprising administering a compoundprovided herein, e.g., a PI3K γ selective inhibitor, a PI3K δ selectiveinhibitor, or a PI3K γ/δ dual inhibitor. Without being limited by aparticular theory, in some embodiments, selectively inhibiting PI3K-γisoform can provide a treatment regimen where adverse effects associatedwith administration of a non-selective PI3K inhibitor are minimized orreduced. Without being limited by a particular theory, in someembodiments, selectively inhibiting PI3K-δ isoform can provide atreatment regimen where adverse effects associated with administrationof a non-selective PI3K inhibitor are minimized or reduced. Withoutbeing limited by a particular theory, in some embodiments, selectivelyinhibiting PI3K-δ and γ isoform can provide a treatment regimen whereadverse effects associated with administration of a non-selective PI3Kinhibitor are minimized or reduced. Without being limited by aparticular theory, it is believed that the adverse effects can bereduced by avoiding the inhibition of other isoforms (e.g., a or β) ofPI3K.

In one embodiment, the adverse effect is hyperglycemia. In anotherembodiment, the adverse effect is rash. In another embodiment, theadverse effect is impaired male fertility that may result frominhibition of β isoform of PI3K (see, e.g., Ciraolo et al., MolecularBiology of the Cell, 21: 704-711 (2010)). In another embodiment, theadverse effect is testicular toxicity that may result from inhibition ofPI3K-β (see, e.g., Wisler et al., Amgen SOT, Abstract ID #2334 (2012)).In another embodiment, the adverse effect is embryonic lethality (see,e.g., Bi et al., J Biol Chem, 274: 10963-10968 (1999)). In anotherembodiment, the adverse effect is defective platelet aggregation (see,e.g., Kulkarni et al., Science, 287: 1049-1053 (2000)). In anotherembodiment, the adverse effect is functionally defective neutrophil(id.).

In certain embodiments, provided herein is a method of treating orpreventing cancer (e.g., colon cancer, melanoma, bladder cancer, renalcancer, breast, lung cancer, glioblastoma, solid tumors, and a cancer ofhematopoietic origin (e.g., DLBCL, CLL, Hodgkin lymphoma, non-Hodgkinlymphomas)) comprising administering to the subject a PI3K inhibitor(e.g., a PI3K-γ inhibitor, e.g., a compound of any of Formulae (I″),(I′), (A′), (I), (A), (II), (III), (IV), (V), (VI), (VII), (IX), (X),(XI), (XII), (XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound 4,as described herein).

Without being bound by a particular theory, a rationale for the use of aPI3K inhibitor to treat or prevent cancer is that cells derived fromtumors (e.g., from CT26 mouse tumors) can suppress anti-tumor immunecell function, including T-cell proliferation, as shown in the examplesprovide herein, and treatment with a compound provided herein canrelease the suppression. The tumor microenvironment can inhibit theactivation and proliferation of immune effector cells due to thepresence of suppressive myeloid cells (e.g., myeloid derived suppressorcells or MDSC and M2 macrophages). Compounds provided herein can affectthe number and activity M2 macrophages in a tumor microenvironment,e.g., reduce or inhibit the level of M2, pro-tumor macrophages. Thereduction or inhibition of M2 macrophages, which produceanti-inflammatory cytokines and other factors, would lead to increasedanti-tumor immunity, including T cell proliferation. Therefore, acompound provided herein can treat or prevent cancer such as coloncancer, melanoma, bladder cancer, renal cancer, breast, lung cancer,glioblastoma, solid tumors, and a cancer of hematopoietic origin (e.g.,lymphoma, DLBCL, CLL, Hodgkin disease, non-Hodgkin lymphomas). Further,it has also been shown in the examples provided herein that anti-PDL1can also release suppression of T cell proliferation by blocking theinteraction between PD1 on T cells and PDL1 on tumor cells andregulatory cells. The cytotoxic T cells that are induced to proliferateand survive by both anti PDL-1 and compound BB are hypothesized to slowtumor growth. Compounds provided herein can relieve immunosuppressionwhich can lead to T cells proliferation and activation. Compoundsprovided herein can treat or prevent cancer by inducing T cell mediatedimmunity. In one embodiment, the compound provided herein can decreasetumor volume. In one embodiment, a combination of a PI3K inhibitor suchas a compound provided herein and anti-PDL1 would be effective intreating or preventing cancer by inducing T cell mediated tumorimmunity. In some embodiments, the effect of a compound provided hereinon T-cell function can be assessed by analyzing the pro-inflammatorycytokine levels in tumor tissues and serum, e.g., a MSD pro-inflammatorypanel. In another embodiment, the pro-inflammatory cytokines areselected from IFN-γ, IL-113, IL-10, IL-12 p70, IL-2, IL-4, IL-5, IL-6,KC/GRO, and TNF-α. In one embodiment, the effect of a compound providedherein on T cell function can be assessed by analyzing the IFN-γ level.For example, tumor tissues and serum treated with a compound providedherein, e.g., Compound 4, can be assessed by analyzing the IFN-γ level.

Treatment of Neuropsychiatric Disorders

In other embodiments, inhibition of PI3K (such as PI3K-δ and/or PI3K-γ)can be used to treat a neuropsychiatric disorder, e.g., an autoimmunebrain disorder. Infectious and immune factors have been implicated inthe pathogenesis of several neuropsychiatric disorders, including, butnot limited to, Sydenham's chorea (SC) (Garvey, M. A. et al. (2005) J.Child Neurol. 20:424-429), Tourette's syndrome (TS), obsessivecompulsive disorder (OCD) (Asbahr, F. R. et al. (1998) Am. J. Psychiatry155:1122-1124), attention deficit/hyperactivity disorder (AD/HD)(Hirschtritt, M. E. et al. (2008) Child Neuropsychol. 1:1-16; Peterson,B. S. et al. (2000) Arch. Gen. Psychiatry 57:364-372), anorexia nervosa(Sokol, M. S. (2000) J. Child Adolesc. Psychopharmacol. 10:133-145;Sokol, M. S. et al. (2002) Am. J. Psychiatry 159:1430-1432), depression(Leslie, D. L. et al. (2008) J. Am. Acad. Child Adolesc. Psychiatry47:1166-1172), and autism spectrum disorders (ASD) (Hollander, E. et al.(1999) Am. J. Psychiatry 156:317-320; Margutti, P. et al. (2006) Curr.Neurovasc. Res. 3:149-157). A subset of childhood obsessive compulsivedisorders and tic disorders has been grouped as Pediatric AutoimmuneNeuropsychiatric Disorders Associated with Streptococci (PANDAS). PANDASdisorders provide an example of disorders where the onset andexacerbation of neuropsychiatric symptoms is preceded by a streptococcalinfection (Kurlan, R., Kaplan, E. L. (2004) Pediatrics 113:883-886;Garvey, M. A. et al. (1998) J. Clin. Neurol. 13:413-423). Many of thePANDAS disorders share a common mechanism of action resulting fromantibody responses against streptococcal associated epitopes, such asGlcNAc, which produces neurological effects (Kirvan. C. A. et al. (2006)J. Neuroimmunol. 179:173-179). Autoantibodies recognizing centralnervous system (CNS) epitopes are also found in sera of most PANDASsubjects (Yaddanapudi, K. et al. (2010) Mol. Psychiatry 15:712-726).Thus, several neuropsychiatric disorders have been associated withimmune and autoimmune components, making them suitable for therapiesthat include PI3K-δ and/or PI3K-γ inhibition.

In certain embodiments, a method of treating (e.g., reducing orameliorating one or more symptoms of) a neuropsychiatric disorder,(e.g., an autoimmune brain disorder), using a PI3K-δ and/or PI3K-γinhibitor is described, alone or in combination therapy. For example,one or more PI3K-δ and/or PI3K-γ inhibitors described herein can be usedalone or in combination with any suitable therapeutic agent and/ormodalities, e.g., dietary supplement, for treatment of neuropsychiatricdisorders. Exemplary neuropsychiatric disorders that can be treated withthe PI3K-δ and/or PI3K-γ inhibitors described herein include, but arenot limited to, PANDAS disorders, Sydenham's chorea, Tourette'ssyndrome, obsessive compulsive disorder, attention deficit/hyperactivitydisorder, anorexia nervosa, depression, and autism spectrum disorders.Pervasive Developmental Disorder (PDD) is an exemplary class of autismspectrum disorders that includes Autistic Disorder, Asperger's Disorder,Childhood Disintegrative Disorder (CDD), Rett's Disorder and PDD-NotOtherwise Specified (PDD-NOS). Animal models for evaluating the activityof the PI3K-δ and/or PI3K-γ inhibitor are known in the art. For example,a mouse model of PANDAS disorders is described in, e.g., Yaddanapudi, K.et al. (2010) supra; and Hoffman, K. I. et al. (2004) J. Neurosci.24:1780-1791.

In some embodiments, provided herein is a method for treating rheumatoidarthritis or asthma in a subject, or for reducing a rheumatoidarthritis-associated symptom or an asthma-associated symptom in asubject, comprising administering an effective amount of a PI3K-γinhibitor to a subject in need thereof, wherein one or more of theadverse effects associated with administration of inhibitors for one ormore other isoforms of PI3K are reduced. In one embodiment, the one ormore other isoforms of PI3K is PI3K-α, PI3K-β, and/or PI3K-δ. In oneembodiment, the one or more other isoforms of PI3K is PI3K-α and/orPI3K-β. In one embodiment, the method is for treating rheumatoidarthritis in a subject, or for reducing a rheumatoidarthritis-associated symptom in a subject. In another embodiment, themethod is for treating asthma in a subject, or for reducing anasthma-associated symptom in a subject.

In some embodiments, provided herein are methods of using a compoundprovided herein, or a pharmaceutically acceptable form (e.g.,pharmaceutically acceptable salts, hydrates, solvates, isomers,prodrugs, and isotopically labeled derivatives) thereof, or apharmaceutical composition as provided herein, to treat diseaseconditions, including, but not limited to, diseases associated withmalfunctioning of one or more types of PI3 kinase. In one embodiment, adetailed description of conditions and disorders mediated by p1108kinase activity is set forth in Sadu et al., WO 01/81346, which isincorporated herein by reference in its entirety for all purposes.

In some embodiments, the disclosure relates to a method of treating ahyperproliferative disorder in a subject that comprises administering tosaid subject a therapeutically effective amount of a compound providedherein, or a pharmaceutically acceptable form (e.g., pharmaceuticallyacceptable salts, hydrates, solvates, isomers, prodrugs, andisotopically labeled derivatives) thereof, or a pharmaceuticalcomposition as provided herein. In some embodiments, said method relatesto the treatment of cancer such as acute myeloid leukemia, thymus,brain, lung, squamous cell, skin, eye, retinoblastoma, intraocularmelanoma, oral cavity and oropharyngeal, bladder, gastric, stomach,pancreatic, bladder, breast, cervical, head, neck, renal, kidney, liver,ovarian, prostate, colorectal, esophageal, testicular, gynecological,thyroid, CNS, PNS, AIDS-related (e.g., Lymphoma and Kaposi's Sarcoma) orviral-induced cancer. In some embodiments, said method relates to thetreatment of a non-cancerous hyperproliferative disorder such as benignhyperplasia of the skin (e.g., psoriasis), restenosis, or prostate(e.g., benign prostatic hypertrophy (BPH)).

Treatment of Cancers

In certain embodiments, provided herein are methods of modulating tumormicroenvironment of cancer cells in a subject, comprising administeringto the subject a therapeutically effective amount of a compound providedherein (e.g., a compound of any of Formulae (I″), (I′), (A′), (I), (A),(II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII),(XIV), (XV), (XVI), and (XVII), e.g., a selective PI3K-γ inhibitor,e.g., Compound 4), or a pharmaceutically acceptable form thereof.

As used herein and unless otherwise specified, “tumor microenvironment”refers to the cellular and extracellular environment where the tumorsare located. This location can include surrounding blood vessels, immunecells, fibroblasts, secreted signaling molecules, and the extracellularmatrix. The tumor microenvironment includes non-neoplastic stromal andimmune cells that provide growth and survival support to the neoplastictumor.

As used herein and unless otherwise specified, “immunotherapy” refers totreatments that stimulate, enhance, or suppress the body's own immunesystem to fight a disease. Diseases that may be suitable forimmunotherapy treatment include, but are not limited to, cancer,inflammatory diseases, and infectious diseases. Immunotherapy includes avariety of treatments that work in different ways. For example, some areintended to boost the immune system defenses in a general way; othershelp train the immune system to recognize and attack cancer cellsspecifically. Cancer immunotherapies include, but are not limited to,cell-based therapies (also known as cancer vaccines), antibodytherapies, and cytokine therapies (e.g., interleukin-2 andinterferon-α).

Many cancers are known to be susceptible to the treatment of one or moreimmunotherapies, including treatment targeting the effector cells in thetumor microenvironment (e.g., immune checkpoint therapy such asPD-1/PD-L1 inhibitors and CTLA-4 inhibitors), treatment targetingsuppressor cells in the tumor microenvironment (e.g., CSF-1R inhibitors(affecting MDSC and TAM) and IDO/TDO inhibitors). Without being limitedby a particular theory, a compound provided herein (e.g., a compound ofany of Formulae (I″), (I′), (A′), (I), (A), (II), (III), (IV), (V),(VI), (VII), (IX), (X), (XI), (XII), (XIII), (XIV), (XV), (XVI), and(XVII), e.g., a selective PI3K-γ inhibitor, e.g., Compound 4) may affectMDSC, TAM, and other components in the tumor microenvironment. The roleof TAM in tumor microenvironment is described, e.g., in Lewis andPollard, Cancer Res. 2006; 66: (2). Jan. 15, 2006.

In one embodiment, the number of one or more pro-tumor immune cells inthe tumor microenvironment is reduced, or the activity of one or morepro-tumor immune cells in the tumor microenvironment is reduced orinhibited, after administration of the compound. In some embodiments,the pro-tumor immune cell is a T-cell, an M2 macrophage, a stromal cell,a dendritic cell, an endothelial cell, or a myeloid cell. In oneembodiment, the myeloid cell is a tumor associated suppressive myeloidcell. In one embodiment, the tumor associated suppressive myeloid cellis identified by (i) CD45+, CD11b+, Ly6C+ and Ly6G+, (ii) CD45+, CD11b+,Ly6C− and Ly6G−, (iii) CD45+, CD11b+, Ly6C− and Ly6G+, or (iv) CD45+,CD11b+, Ly6C+ and Ly6G−. In one embodiment, the tumor associatedsuppressive myeloid cell is a tumor associated macrophage (TAM), amyeloid derived suppressor cell (MDSC), a monocytic immature myeloidcell (iMc), or a granulocytic iMc/neutrophil. In one embodiment, the TAMis identified by CD45+, CD11b+, Ly6C−, and Ly6G−. In one embodiment, themyeloid derived suppressor cell (MDSC) is identified by CD45+, CD11b+,Ly6C− and Ly6G+. In one embodiment, the monocytic immature myeloid cell(iMc) is identified by CD45+, CD11b+, Ly6C+ and Ly6G−. In oneembodiment, the granulocytic iMc/neutrophil is identified by CD45+,CD11b+, Ly6C+ and Ly6G+. See e.g., Coussens L M. et al., Cancer Discov.2011 June; 1(1):54-67.

In one embodiment, the activation of M2 macrophage in the tumormicroenvironment is reduced or inhibited after administration of thecompound. In one embodiment, the p-AKT level in the M2 macrophage isreduced after administration of the compound. In one embodiment, thenumber of M2 macrophage cells in the tumor microenvironment is reducedafter administration of the compound. In one embodiment, the migrationof M2 macrophage cells into the tumor microenvironment is reduced orinhibited after administration of the compound. In one embodiment, thedifferentiation of myeloid cells into M2 macrophage cells in the tumormicroenvironment is reduced or inhibited after administration of thecompound. In one embodiment, the differentiation into M2 macrophagecells is measured by Arginase-1 (ARG1) level or VEGF level, and the ARG1level or VEGF level is reduced by at least 10%, 20%, 30%, 40%, 50%, 60%,70%, 80%, or 90% compared to a reference value.

In one embodiment, the number of myeloid-derived suppressor cells in thetumor microenvironment is reduced after administration of the compound.In one embodiment, the differentiation of bone marrow cells intomyeloid-derived suppressor cells is reduced or inhibited afteradministration of the compound. In one embodiment, the differentiationinto myeloid-derived suppressor cells is measured by Arginase-1 (ARG1)level, VEGF level, or iNOS level, and the ARG1 level, VEGF level, oriNOS level is reduced by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%,80%, or 90% compared to a reference value.

In one embodiment, the production of proangiogeneic factor is reduced orinhibited after administration of the compound. In one embodiment, theproangiogeneic factor is reduced or inhibited by reduction or inhibitionof macrophage or MDSC differentiation. In one embodiment, theproangiogeneic factor is VEGF.

In one embodiment, the effect of the compounds provided herein (e.g.,Compound 4) on MDSC (e.g., human MDSC) function is measured byexpression of iNOS and arginase and production of ROS and IL-10,measured by the suppression function of the MDSC (e.g., in co-cultureassays with CD8+), measured by activation of pAKT in response to astimulant (e.g., CXCL12, IL-1b, TNF-α, or CSF1), or measured bytranswell chemotaxis assays (T cells and MDSC).

In one embodiment, the effect of the compounds provided herein (e.g.,Compound 4) on MDSC (e.g., murine MDSC) function and macrophageM2-polarization is measured by isolating myeloid cells from bone marrow,polarizing with IFNg or IL-4 and then testing for secretion of TNF-α,IL-12, ROS production in M1 and IL-10, IL-1b, or VEGF, or measured bymethods provided herein or elsewhere.

In one embodiment, the effect of the compounds provided herein (e.g.,Compound 4) on myeloid and CD8+ is measured by in vivo models (e.g.,MC38 and 4T1). In one embodiment, the effect is measured by TGI, MDSCand macrophage infiltrate, CD8+, and IFN-gamma production in CD8+.

In one embodiment, the effect of the compounds provided herein (e.g.,Compound 4) on myeloid and CD8+ is measured by QT-PCR or intracellularFACS of myeloid infiltrate. In one embodiment, the effect is measured byexpression of functional markers (e.g., iNOS, arginase, or IL-10).

In one embodiment, the number of one or more anti-tumor immune cells inthe tumor microenvironment is increased, or the activity of one or moreanti-tumor immune cells in the tumor microenvironment is increased,after administration of the compound.

In one embodiment, the cancer susceptible to the treatment of one ormore immunotherapies is a hematological cancer. In one embodiment, thehematological cancer is chronic lymphocytic leukemia (CLL). In oneembodiment, the tumor microenvironment is a CLL proliferation center. Inone embodiment, the hematological cancer is lymphoma.

In one embodiment, the cancer susceptible to the treatment of one ormore immunotherapies is a solid tumor. In one embodiment, the solidtumor is lung cancer, breast cancer, colon cancer, or glioblastoma. Inone embodiment, the cancer is selected from one or more of: a cancer ofthe pulmonary system, a brain cancer, a cancer of the gastrointestinaltract, a skin cancer, a genitourinary cancer, a pancreatic cancer, alung cancer, a medulloblastoma, a basal cell carcinoma, a glioma, abreast cancer, a prostate cancer, a testicular cancer, an esophagealcancer, a hepatocellular cancer, a gastric cancer, a gastrointestinalstromal tumor (GIST), a colon cancer, a colorectal cancer, an ovariancancer, a melanoma, a neuroectodermal tumor, head and neck cancer, asarcoma, a soft-tissue sarcoma, fibrosarcoma, myxosarcoma, liposarcoma,a chondrosarcoma, an osteogenic sarcoma, a chordoma, an angiosarcoma, anendotheliosarcoma, a lymphangiosarcoma, a lymphangioendotheliosarcoma, asynovioma, a mesothelioma, a leiomyosarcoma, a cervical cancer, auterine cancer, an endometrial cancer, a carcinoma, a bladder carcinoma,an epithelial carcinoma, a squamous cell carcinoma, an adenocarcinoma, abronchogenic carcinoma, a renal cell carcinoma, a hepatoma, a bile ductcarcinoma, a neuroendocrine cancer, a carcinoid tumor, diffuse typegiant cell tumor, andglioblastoma. In one embodiment, the solid tumor ismelanoma, bladder cancer, head and neck cancer, lung cancer (e.g.,non-small cell lung cancer), renal cell carcinoma, ovarian cancer,breast cancer (e.g., triple-negative breast cancer), colon cancer, orglioblastoma.

In one embodiment, the solid tumor is melanoma. In one embodiment, thesolid tumor is lung cancer. In one embodiment, the solid tumor isnon-small cell lung cancer. In one embodiment, the solid tumor is renalcell carcinoma. Melanoma, lung cancer (e.g., non-small cell lungcancer), and renal cell carcinoma are known to be sensitive toimmunotherapies. Data linking a poor prognosis to high TAM cell countshave been reported in breast, prostate, endometrial, bladder, kidney,esophageal, superficial, carcinoma, melanoma, and follicular lymphomacancers. See e.g., Lewis and Pollard, Cancer Res. 2006; 66: (2). Jan.15, 2006. One anti-PD-1 antibody drug, nivolumab, (Opdivo-Bristol MyersSquibb), produced complete or partial responses in non-small-cell lungcancer, melanoma, and renal-cell cancer, in a clinical trial with atotal of 296 patients.

In one embodiment, the solid tumor is head and neck cancer. Head andneck tumors tend to be highly immunogenic and have stronganti-PD-1/PD-L1 efficacy. In one embodiment, the solid tumor is bladdercancer. Bladder cancer also has strong anti-PD-1/PD-L1 efficacy. A highnumber of TAM cells has been associated with a poor prognosis andincreased tumor angiogenesis in bladder cancer.

In one embodiment, the solid tumor is breast cancer. In one embodiment,the breast cancer is triple-negative breast cancer. A high number of TAMcells has been associated with a poor prognosis of breast cancer. Seee.g., Lewis and Pollard, Cancer Res. 2006; 66: (2). Jan. 15, 2006. Inone embodiment, the solid tumor is ovarian cancer. In one embodiment,the solid tumor is colon cancer. Breast cancer, ovarian cancer, andcolon cancer are known to be sensitive to immunotherapies (e.g.,bevacizumab and trastuzumab) and can also have anti-PD-1/PD-L1 efficacy.

In one embodiment, the solid tumor is glioblastoma. In one embodiment,the solid tumor is glioblastoma multiforme. It has been reported thatPI3K-gamma expression is upregulated in brain microglia. Without beinglimited by a particular theory, PI3K-γ inhibitors provided herein (e.g.,Compound BB) may have P-glycoprotein inhibitory activity and thus cancross the blood brain barrier.

In one embodiment, the anti-tumor immune attack by effector T cells isincreased, vascularization of the tumor is reduced, extracellular matrix(ECM) breakdown is reduced, or tumor growth is decreased, compared to areference value, after administration of the compound.

In one embodiment, the tumor volume of the cancer is reduced afteradministration of the compound. In one embodiment, the tumor volume ofthe cancer is reduced by at least 10%, 20%, 30%, 50%, 60%, or 60%,compared to a reference value.

In one embodiment, the level of apoptosis of the cancer cells isincreased after administration of the compound. In one embodiment, thelevel of apoptosis of the cancer cells is increased by at least 10%,20%, 30%, 40%, or 50%, compared to a reference value.

In some embodiments, the subject is naive to immunotherapy treatment. Insome embodiments, the subject is naive to radiation therapy treatment.In some embodiments, the subject is naive to chemotherapy treatment.

In some embodiments, the subject has been pre-treated or previouslytreated with one or more immunotherapy treatments. In one embodiment,the subject is responsive to the pre-treatment or previous treatmentwith the immunotherapy. In one embodiment, the immunotherapy treatmentis a checkpoint treatment such as a PD-1 or PD-L1 inhibitor. In oneembodiment, the subject is a smoker. It has been reported that smokerpatients may respond better to immunotherapy (e.g., a PD-L1 inhibitorMPDL3280A) than non-smoker patients in a phase I clinical study forpatients with melanoma or cancers of the lung, kidney, colon, GI tract,or head and neck cancers.

In one embodiment, the cancer is melanoma, and the subject has beenpre-treated or previously treated with one or more immunotherapytreatments. In one embodiment, the subject has been pre-treated orpreviously treated with two or more immunotherapy treatments.

In one embodiment, the cancer is head and neck cancer, lung cancer(e.g., non-small cell lung cancer), renal cell carcinoma, or bladdercancer, and the subject has been pre-treated or previously treated withone immunotherapy treatment.

In one embodiment, the cancer is breast cancer (e.g., triple-negativebreast cancer), ovarian cancer, glioblastoma, or colon cancer, and thesubject is naive to immunotherapy treatment.

In one embodiment, provided herein is a method of treating, preventing,or managing melanoma in a subject, comprising administering to thesubject a therapeutically effective amount of a compound provided herein(e.g., Compound 4), or a pharmaceutically acceptable form thereof,wherein the subject has been pre-treated or previously treated with oneor more immunotherapy treatments. In one embodiment, the immunotherapytreatment is ipilimumab (Yervoy), interleukin-2, vemurafenib,dabrafenib, or trametinib.

In one embodiment, provided herein is a method of treating, preventing,or managing lung cancer (e.g., non-small cell lung cancer) in a subject,comprising administering to the subject a therapeutically effectiveamount of a compound provided herein (e.g., Compound 4), or apharmaceutically acceptable form thereof, wherein the subject has beenpre-treated or previously treated with one or more immunotherapytreatments. In one embodiment, the immunotherapy treatment isbevacizumab, erlotinib, gefitinib, afatinib, or denosumab.

In one embodiment, provided herein is a method of treating, preventing,or managing renal cell carcinoma in a subject, comprising administeringto the subject a therapeutically effective amount of a compound providedherein (e.g., Compound 4), or a pharmaceutically acceptable formthereof, wherein the subject has been pre-treated or previously treatedwith one or more immunotherapy treatments. In one embodiment, theimmunotherapy treatment is bevacizumab, interleukin-2, axitinib,carfilzomib, everolimus, interferon-α, lenalidomide, pazopanib,sirolimus (rapamycin), sorafenib, sunitinib, temsirolimus, thalidomide,or tivozanib.

In one embodiment, provided herein is a method of treating, preventing,or managing bladder cancer in a subject, comprising administering to thesubject a therapeutically effective amount of a compound provided herein(e.g., Compound 4), or a pharmaceutically acceptable form thereof,wherein the subject has been pre-treated or previously treated with oneor more immunotherapy treatments. In one embodiment, the immunotherapytreatment is Bacillus Calmette-Guérin (BCG).

In one embodiment, provided herein is a method of treating, preventing,or managing head and neck cancer in a subject, comprising administeringto the subject a therapeutically effective amount of a compound providedherein (e.g., Compound 4), or a pharmaceutically acceptable formthereof, wherein the subject has been pre-treated or previously treatedwith one or more immunotherapy treatments. In one embodiment, theimmunotherapy treatment is cetuximab, nimotuzumab, bevacizumab, orerlotinib.

In one embodiment, provided herein is a method of treating, preventing,or managing breast cancer (e.g., triple-negative breast cancer) in asubject, comprising administering to the subject a therapeuticallyeffective amount of a compound provided herein (e.g., Compound 4), or apharmaceutically acceptable form thereof, wherein the subject has beenpre-treated or previously treated with one or more immunotherapytreatments. In one embodiment, the immunotherapy treatment isbevacizumab or trastuzumab.

In one embodiment, provided herein is a method of treating, preventing,or managing ovarian cancer in a subject, comprising administering to thesubject a therapeutically effective amount of a compound provided herein(e.g., Compound 4), or a pharmaceutically acceptable form thereof,wherein the subject has been pre-treated or previously treated with oneor more immunotherapy treatments. In one embodiment, the immunotherapytreatment is bevacizumab.

In one embodiment, provided herein is a method of treating, preventing,or managing colon cancer in a subject, comprising administering to thesubject a therapeutically effective amount of a compound provided herein(e.g., Compound 4), or a pharmaceutically acceptable form thereof,wherein the subject has been pre-treated or previously treated with oneor more immunotherapy treatments. In one embodiment, the immunotherapytreatment is bevacizumab, cetuximab, or panitumumab.

In some embodiments, the disclosure relates to a method of treating acancer of hematopoietic origin. In certain embodiments, the cancer ofhematopoietic origin is lymphoma or leukemia. In some embodiments, thecancer of hematopoietic origin is selected from acute lymphocyticleukemia (ALL), which includes B-lineage ALL and T-lineage ALL, chroniclymphocytic leukemia (CLL), prolymphocytic leukemia (PLL), hairy cellleukemia (HLL) and Waldenstrom's macroglobulinemia (WM); peripheral Tcell lymphomas (PTCL), adult T cell leukemia/lymphoma (ATLL), cutaneousT-cell lymphoma (CTCL), large granular lymphocytic leukemia (LGL), acutemyelocytic leukemia (AML), Hodgkin lymphoma (HL), non-Hodgkin lymphoma(NHL), follicular lymphoma, diffuse large B-cell lymphoma (DLBCL),mantle cell lymphoma (MCL), mastocytosis (e.g., systemic mastocytosis),multiple myeloma (MM), myelodysplastic syndrome (MDS),myeloproliferative disorder (MPD) (e.g., polycythemia Vera (PV),essential thrombocytosis (ET), agnogenic myeloid metaplasia (AMM) a.k.a.myelofibrosis (MF), chronic idiopathic myelofibrosis, chronicmyelogenous leukemia (CML), chronic neutrophilic leukemia (CNL), andhypereosinophilic syndrome (HES)).

In some embodiments, the disclosure relates to a method of treating asolid tumor. In some embodiments, the solid tumor is selected fromovarian cancer, colon cancer, fibrosarcoma, pancreatic cancer, lungcancer, breast cancer, lymphoma, melanoma, and glioblastoma. In someembodiment, the solid tumor is a CNS tumor. In one embodiment, the CNStumor is glioblastoma. The ADME data provide herein indicate that acompound provide herein (e.g., Compound BB) may show good permeabilitycross blood-brain-barrier and can achieving efficacious concentration ina CNS tumor.

In one embodiment, a PI3K-γ inhibitor such a Compound BB can be aninhibitor of P-gp (P-glycoprotein). P-glycoprotein impedes the entry ofvarious drugs that are used in the treatment, for example, of centralnervous system diseases. Without being bound by a particular theory, theP-gp substrate may help maintain normal levels of P-gp activity in apatient being treated with a PI3K-γ inhibitor. In some embodiments, aPI3K-γ inhibitor such a Compound BB may not be effluxed from a tumor andthus can maintain efficacious concentration of the a PI3K-γ inhibitor ina tumor. For example, the concentration can be maintained for about atleast 6 hours, at least 10 hours, at least 12 hours, at least 24 hours,or at least 48 hours. In one embodiment, a PI3K-γ inhibitor such aCompound BB can be administered once daily.

In some embodiments, a PI3K-γ inhibitor such a Compound BB isadministered to a patient in combination with a second therapeutic thatis a P-gp substrate. In another embodiment, a PI3K-γ inhibitor such aCompound BB can inhibit the efflux of the second therapeutic such as acancer drug that is a P-gp substrate. Therefore, a PI3K-γ inhibitorprovided herein such a Compound BB can be efficacious in maintaining theconcentration of the co-administered cancer drug in a tumor. Forexample, the concentration can be maintained for about at least 6 hours,at least 10 hours, at least 12 hours, at least 24 hours, or at least 48hours. In one embodiment, a PI3K-γ inhibitor such a Compound BB can beadministered once daily.

P-glycoprotein is a component of the blood-brain barrier and is presenton the surface of the endothelial cells of the barrier. A PI3K-γinhibitor provided herein such as Compound BB can be a P-glycoproteininhibitor and thus can cross the blood brain barrier. In someembodiments, a PI3K-γ inhibitor provided herein such as Compound BB canmaintain an efficacious concentration in CNS tumor or a brain tumor(e.g., glioblastoma).

As used herein “solid tumor” refers to an abnormal mass of tissue. Solidtumors may be benign or malignant. A solid tumor grows in an anatomicalsite outside the bloodstream (in contrast, for example, to cancers ofhematopoietic origin such as leukemias) and requires the formation ofsmall blood vessels and capillaries to supply nutrients, etc. to thegrowing tumor mass. Solid tumors are named for the type of cells thatform them. Non-limiting examples of solid tumors are sarcomas,carcinomas (epithelial tumors), melanomas, and glioblastomas.

In some embodiments, the disclosure relates to a method of inhibitinggrowth of a tumor. “Inhibiting growth of a tumor” refers to slowingtumor growth and/or reducing tumor size. “Inhibiting growth of a tumor”thus includes killing tumor cells as well as slowing or arresting tumorcell growth.

Exemplary solid tumors include, but are not limited to, biliary cancer(e.g., cholangiocarcinoma), bladder cancer, breast cancer (e.g.,adenocarcinoma of the breast, papillary carcinoma of the breast, mammarycancer, medullary carcinoma of the breast), brain cancer (e.g.,meningioma; glioma, e.g., astrocytoma, oligodendroglioma;medulloblastoma), cervical cancer (e.g., cervical adenocarcinoma),colorectal cancer (e.g., colon cancer, rectal cancer, colorectaladenocarcinoma), gastric cancer (e.g., stomach adenocarcinoma),gastrointestinal stromal tumor (GIST), head and neck cancer (e.g., headand neck squamous cell carcinoma, oral cancer (e.g., oral squamous cellcarcinoma (OSCC)), kidney cancer (e.g., nephroblastoma a.k.a. Wilms'tumor, renal cell carcinoma), liver cancer (e.g., hepatocellular cancer(HCC), malignant hepatoma), lung cancer (e.g., bronchogenic carcinoma,small cell lung cancer (SCLC), non-small cell lung cancer (NSCLC),adenocarcinoma of the lung), neuroblastoma, neurofibroma (e.g.,neurofibromatosis (NF) type 1 or type 2, schwannomatosis),neuroendocrine cancer (e.g., gastroenteropancreatic neuroendoctrinetumor (GEP-NET), carcinoid tumor), osteosarcoma, ovarian cancer (e.g.,cystadenocarcinoma, ovarian embryonal carcinoma, ovarianadenocarcinoma), pancreatic cancer (e.g., pancreatic andenocarcinoma,intraductal papillary mucinous neoplasm (IPMN)), prostate cancer (e.g.,prostate adenocarcinoma), skin cancer (e.g., squamous cell carcinoma(SCC), keratoacanthoma (A), melanoma, basal cell carcinoma (BCC)) andsoft tissue sarcoma (e.g., malignant fibrous histiocytoma (MFH),liposarcoma, malignant peripheral nerve sheath tumor (MPNST),chondrosarcoma, fibrosarcoma, myxosarcoma, osteosarcoma).

In some embodiments, the disclosure relates to a method of treating acancer of hematopoietic origin comprising administering to a subject agamma selective compound. In certain embodiments, the cancer ofhematopoietic origin is lymphoma or leukemia. In some embodiments, thecancer of hematopoietic origin is selected from acute lymphocyticleukemia (ALL), which includes B-lineage ALL and T-lineage ALL, chroniclymphocytic leukemia (CLL), prolymphocytic leukemia (PLL), hairy cellleukemia (HLL) and Waldenstrom's macroglobulinemia (WM); peripheral Tcell lymphomas (PTCL), adult T cell leukemia/lymphoma (ATLL), cutaneousT-cell lymphoma (CTCL), large granular lymphocytic leukemia (LGL), acutemyelocytic leukemia (AML), Hodgkin lymphoma (HL), non-Hodgkin lymphoma(NHL), follicular lymphoma, diffuse large B-cell lymphoma (DLBCL),mantle cell lymphoma (MCL), mastocytosis (e.g., systemic mastocytosis),multiple myeloma (MM), myelodysplastic syndrome (MDS),myeloproliferative disorder (MPD) (e.g., polycythemia Vera (PV),essential thrombocytosis (ET), agnogenic myeloid metaplasia (AMM) a.k.a.myelofibrosis (MF), chronic idiopathic myelofibrosis, chronicmyelogenous leukemia (CML), chronic neutrophilic leukemia (CNL), andhypereosinophilic syndrome (HES)).

In some embodiments, the disclosure relates to a method of treating asolid tumor comprising administering to a subject a gamma selectivecompound. In some embodiments, the solid tumor is selected from ovariancancer, colon cancer, fibrosarcoma, pancreatic cancer, lung cancer,breast cancer, lymphoma, melanoma, and glioblastoma.

In some embodiments, the disclosure relates to a method of treating aninflammatory disease comprising administering to a subject a gammaselective compound.

In some embodiment, the gamma selective compound has a delta/gammaselectivity ratio of >1 to <10, 10 to <50, or 50 to <350 can be combinedwith a compound that has a gamma/delta selectivity ratio of greater thana factor of about 1, greater than a factor of about 2, greater than afactor of about 3, greater than a factor of about 5, greater than afactor of about 10, greater than a factor of about 50, greater than afactor of about 100, greater than a factor of about 200, greater than afactor of about 400, greater than a factor of about 600, greater than afactor of about 800, greater than a factor of about 1000, greater than afactor of about 1500, greater than a factor of about 2000, greater thana factor of about 5000, greater than a factor of about 10,000, orgreater than a factor of about 20,000.

Patients that can be treated with a compound provided herein, or apharmaceutically acceptable form (e.g., pharmaceutically acceptablesalts, hydrates, solvates, isomers, prodrugs, and isotopically labeledderivatives) thereof, or a pharmaceutical composition as providedherein, according to the methods as provided herein include, forexample, but not limited to, patients that have been diagnosed as havingpsoriasis; restenosis; atherosclerosis; ischemic stroke, BPH; breastcancer such as a ductal carcinoma, lobular carcinoma, medullarycarcinomas, colloid carcinomas, tubular carcinomas, and inflammatorybreast cancer; ovarian cancer, including epithelial ovarian tumors suchas adenocarcinoma in the ovary and an adenocarcinoma that has migratedfrom the ovary into the abdominal cavity; uterine cancer; cervicalcancer such as adenocarcinoma in the cervix epithelial includingsquamous cell carcinoma and adenocarcinomas; prostate cancer, such as aprostate cancer selected from the following: an adenocarcinoma or anadenocarcinoma that has migrated to the bone; pancreatic cancer such asepitheliod carcinoma in the pancreatic duct tissue and an adenocarcinomain a pancreatic duct; bladder cancer such as a transitional cellcarcinoma in urinary bladder, urothelial carcinomas (transitional cellcarcinomas), tumors in the urothelial cells that line the bladder,squamous cell carcinomas, adenocarcinomas, and small cell cancers;leukemia such as acute lymphoblastic leukemia, chronic myelogenousleukemia, hairy cell leukemia, myelodysplasia, myeloproliferativedisorders, NK cell leukemia (e.g., blastic plasmacytoid dendritic cellneoplasm), acute myelogenous leukemia (AML), chronic myelogenousleukemia (CML), mastocytosis, chronic lymphocytic leukemia (CLL),multiple myeloma (MM), and myelodysplastic syndrome (MDS); bone cancer;lung cancer such as non-small cell lung cancer (NSCLC), which is dividedinto squamous cell carcinomas, adenocarcinomas, and large cellundifferentiated carcinomas, and small cell lung cancer; skin cancersuch as basal cell carcinoma, melanoma, squamous cell carcinoma andactinic keratosis, which is a skin condition that sometimes developsinto squamous cell carcinoma; eye retinoblastoma; cutaneous orintraocular (eye) melanoma; primary liver cancer; kidney cancer; thyroidcancer such as papillary, follicular, medullary and anaplastic; lymphomasuch as diffuse large B-cell lymphoma, B-cell immunoblastic lymphoma, NKcell lymphoma (e.g., blastic plasmacytoid dendritic cell neoplasm), andBurkitt lymphoma; Kaposi's Sarcoma; viral-induced cancers includinghepatitis B virus (HBV), hepatitis C virus (HCV), and hepatocellularcarcinoma; human lymphotropic virus-type 1 (HTLV-1) and adult T-cellleukemia/lymphoma; and human papilloma virus (HPV) and cervical cancer;central nervous system cancers (CNS) such as primary brain tumor, whichincludes gliomas (astrocytoma, anaplastic astrocytoma, or glioblastomamultiforme), oligodendroglioma, ependymoma, meningioma, lymphoma,schwannoma, and medulloblastoma; peripheral nervous system (PNS) cancerssuch as acoustic neuromas and malignant peripheral nerve sheath tumor(MPNST) including neurofibromas and schwannomas, malignant fibrocytoma,malignant fibrous histiocytoma, malignant meningioma, malignantmesothelioma, and malignant mixed Müllerian tumor; oral cavity andoropharyngeal cancers such as, hypopharyngeal cancer, laryngeal cancer,nasopharyngeal cancer, and oropharyngeal cancer; stomach cancers such aslymphomas, gastric stromal tumors, and carcinoid tumors; testicularcancers such as germ cell tumors (GCTs), which include seminomas andnonseminomas, and gonadal stromal tumors, which include Leydig celltumors and Sertoli cell tumors; thymus cancer such as to thymomas,thymic carcinomas, Hodgkin lymphoma, non-Hodgkin lymphomas, carcinoidsor carcinoid tumors; rectal cancer; and colon cancer.

Patients that can be treated with compounds provided herein, orpharmaceutically acceptable salt, ester, prodrug, solvate, hydrate orderivative of said compounds, according to the methods provided hereininclude, for example, patients that have been diagnosed as havingconditions including, but not limited to, acoustic neuroma,adenocarcinoma, adrenal gland cancer, anal cancer, angiosarcoma (e.g.,lymphangiosarcoma, lymphangioendotheliosarcoma, hemangiosarcoma), benignmonoclonal gammopathy, biliary cancer (e.g., cholangiocarcinoma),bladder cancer, breast cancer (e.g., adenocarcinoma of the breast,papillary carcinoma of the breast, mammary cancer, medullary carcinomaof the breast), brain cancer (e.g., meningioma; glioma, e.g.,astrocytoma, oligodendroglioma; medulloblastoma), bronchus cancer,cervical cancer (e.g., cervical adenocarcinoma), choriocarcinoma,chordoma, craniopharyngioma, colorectal cancer (e.g., colon cancer,rectal cancer, colorectal adenocarcinoma), epithelial carcinoma,ependymoma, endotheliosarcoma (e.g., Kaposi's sarcoma, multipleidiopathic hemorrhagic sarcoma), endometrial cancer, esophageal cancer(e.g., adenocarcinoma of the esophagus, Barrett's adenocarinoma), Ewingsarcoma, familiar hypereosinophilia, gastric cancer (e.g., stomachadenocarcinoma), gastrointestinal stromal tumor (GIST), head and neckcancer (e.g., head and neck squamous cell carcinoma, oral cancer (e.g.,oral squamous cell carcinoma (OSCC)), heavy chain disease (e.g., alphachain disease, gamma chain disease, mu chain disease), hemangioblastoma,inflammatory myofibroblastic tumors, immunocytic amyloidosis, kidneycancer (e.g., nephroblastoma a.k.a. Wilms' tumor, renal cell carcinoma),liver cancer (e.g., hepatocellular cancer (HCC), malignant hepatoma),lung cancer (e.g., bronchogenic carcinoma, small cell lung cancer(SCLC), non-small cell lung cancer (NSCLC), adenocarcinoma of the lung),leukemia (e.g., acute lymphoblastic leukemia (ALL), which includesB-lineage ALL and T-lineage ALL, chronic lymphocytic leukemia (CLL),prolymphocytic leukemia (PLL), hairy cell leukemia (HLL) andWaldenstrom's macroglobulinemia (WM); peripheral T cell lymphomas(PTCL), adult T cell leukemia/lymphoma (ATLL), cutaneous T-cell lymphoma(CTCL), large granular lymphocytic leukemia (LGL), acute myelocyticleukemia (AML), chronic myelocytic leukemia (CML), chronic lymphocyticleukemia (CLL)), lymphoma (e.g., Hodgkin lymphoma (HL), non-Hodgkinlymphoma (NHL), follicular lymphoma, diffuse large B-cell lymphoma(DLBCL), mantle cell lymphoma (MCL)), leiomyosarcoma (LMS), mastocytosis(e.g., systemic mastocytosis), multiple myeloma (MM), myelodysplasticsyndrome (MDS), mesothelioma, myeloproliferative disorder (MPD) (e.g.,polycythemia Vera (PV), essential thrombocytosis (ET), agnogenic myeloidmetaplasia (AMM) a.k.a. myelofibrosis (MF), chronic idiopathicmyelofibrosis, chronic myelogenous leukemia (CML), chronic neutrophilicleukemia (CNL), hypereosinophilic syndrome (HES)), neuroblastoma,neurofibroma (e.g., neurofibromatosis (NF) type 1 or type 2,schwannomatosis), neuroendocrine cancer (e.g., gastroenteropancreaticneuroendoctrine tumor (GEP-NET), carcinoid tumor), osteosarcoma, ovariancancer (e.g., cystadenocarcinoma, ovarian embryonal carcinoma, ovarianadenocarcinoma), Paget's disease of the vulva, Paget's disease of thepenis, papillary adenocarcinoma, pancreatic cancer (e.g., pancreaticandenocarcinoma, intraductal papillary mucinous neoplasm (IPMN)),pinealoma, primitive neuroectodermal tumor (PNT), prostate cancer (e.g.,prostate adenocarcinoma), rhabdomyosarcoma, retinoblastoma, salivarygland cancer, skin cancer (e.g., squamous cell carcinoma (SCC),keratoacanthoma (KA), melanoma, basal cell carcinoma (BCC)), small bowelcancer (e.g., appendix cancer), soft tissue sarcoma (e.g., malignantfibrous histiocytoma (MFH), liposarcoma, malignant peripheral nervesheath tumor (MPNST), chondrosarcoma, fibrosarcoma, myxosarcoma),sebaceous gland carcinoma, sweat gland carcinoma, synovioma, testicularcancer (e.g., seminoma, testicular embryonal carcinoma), thyroid cancer(e.g., papillary carcinoma of the thyroid, papillary thyroid carcinoma(PTC), medullary thyroid cancer), and Waldenstrom's macroglobulinemia.

Without being limited by a particular theory, in one embodiment, thecancer or disease being treated or prevented, such as a blood disorderor hematologic malignancy, has a high expression level of one or morePI3K isoform(s) (e.g., PI3K-α, PI3K-β, PI3K-δ, or PI3K-γ, or acombination thereof). In one embodiment, the cancer or disease that canbe treated or prevented by methods, compositions, or kits providedherein includes a blood disorder or a hematologic malignancy, including,but not limited to, myeloid disorder, lymphoid disorder, leukemia,lymphoma, myelodysplastic syndrome (MDS), myeloproliferative disease(MPD), mast cell disorder, and myeloma (e.g., multiple myeloma), amongothers. In one embodiment, the blood disorder or the hematologicmalignancy includes, but is not limited to, acute lymphoblastic leukemia(ALL), T-cell ALL (T-ALL), B-cell ALL (B-ALL), acute myeloid leukemia(AML), chronic lymphocytic leukemia (CLL), chronic myelogenous leukemia(CML), blast phase CML, small lymphocytic lymphoma (SLL), CLL/SLL,transformed CLL, Richter syndrome Hodgkin lymphoma (HL), non-Hodgkinlymphoma (NHL), B-cell NHL, T-cell NHL, indolent NHL (iNHL), diffuselarge B-cell lymphoma (DLBCL), mantle cell lymphoma (MCL), aggressiveB-cell NHL, B-cell lymphoma (BCL), Richter's syndrome (RS), T-celllymphoma (TCL), peripheral T-cell lymphoma (PTCL), cutaneous T-celllymphoma (CTCL), transformed mycosis fungoides, Sézary syndrome,anaplastic large-cell lymphoma (ALCL), follicular lymphoma (FL),Waldenstrom macroglobulinemia (WM), lymphoplasmacytic lymphoma, Burkittlymphoma, multiple myeloma (MM), amyloidosis, MPD, essentialthrombocytosis (ET), myelofibrosis (MF), polycythemia vera (PV), chronicmyelomonocytic leukemia (CMML), myelodysplastic syndrome (MDS),angioimmunoblastic lymphoma, high-risk MDS, and low-risk MDS. In oneembodiment, the hematologic malignancy is relapsed. In one embodiment,the hematologic malignancy is refractory. In one embodiment, the canceror disease is in a pediatric patient (including an infantile patient).In one embodiment, the cancer or disease is in an adult patient.Additional embodiments of a cancer or disease being treated or preventedby methods, compositions, or kits provided herein are described hereinelsewhere.

In exemplary embodiments, the cancer or hematologic malignancy is CLL.In exemplary embodiments, the cancer or hematologic malignancy isCLL/SLL. In exemplary embodiments, the cancer or hematologic malignancyis transformed CLL or Richter syndrome. In exemplary embodiments, thecancer or hematologic malignancy is SLL. In one embodiment, withoutbeing limited by a particular theory, a compound provided herein (e.g.,a PI3K-γ selective compound provided herein) inhibits the migrationand/or activation of T-cells and myeloid cells (e.g., macrophages orpolarized M2 macrophages), reducing survival and/or proliferativesupport provided by those cells to malignant CLL cells within the tumormicroenvironment (TME). In one embodiment, without being limited by aparticular theory, the migration of CD3+ T cells to the CLL-associatedchemokine CXCL12 is blocked by a compound provided herein (e.g., aPI3K-γ selective compound provided herein). In another embodiment,without being limited by a particular theory, a compound provided herein(e.g., a PI3K-γ selective compound provided herein) block the myeloidcell mediated re-growth of a cancer following chemotherapy through itseffects on inhibiting the post-chemotherapy migration of myeloid cellsinto a tumor.

In exemplary embodiments, the cancer or hematologic malignancy is iNHL.In exemplary embodiments, the cancer or hematologic malignancy is DLBCL.In exemplary embodiments, the cancer or hematologic malignancy is B-cellNHL (e.g., aggressive B-cell NHL). In exemplary embodiments, the canceror hematologic malignancy is MCL. In exemplary embodiments, the canceror hematologic malignancy is RS. In exemplary embodiments, the cancer orhematologic malignancy is AML. In exemplary embodiments, the cancer orhematologic malignancy is MM. In exemplary embodiments, the cancer orhematologic malignancy is ALL. In exemplary embodiments, the cancer orhematologic malignancy is T-ALL. In exemplary embodiments, the cancer orhematologic malignancy is B-ALL. In exemplary embodiments, the cancer orhematologic malignancy is TCL. In exemplary embodiments, the cancer orhematologic malignancy is ALCL. In exemplary embodiments, the cancer orhematologic malignancy is leukemia. In exemplary embodiments, the canceror hematologic malignancy is lymphoma. In exemplary embodiments, thecancer or hematologic malignancy is T-cell lymphoma. In exemplaryembodiments, the cancer or hematologic malignancy is MDS (e.g., lowgrade MDS). In exemplary embodiments, the cancer or hematologicmalignancy is MPD. In exemplary embodiments, the cancer or hematologicmalignancy is a mast cell disorder. In exemplary embodiments, the canceror hematologic malignancy is Hodgkin lymphoma (HL). In exemplaryembodiments, the cancer or hematologic malignancy is non-Hodgkinlymphoma. In exemplary embodiments, the cancer or hematologic malignancyis PTCL. In exemplary embodiments, the cancer or hematologic malignancyis CTCL (e.g., mycosis fungoides or Sézary syndrome). In exemplaryembodiments, the cancer or hematologic malignancy is WM. In exemplaryembodiments, the cancer or hematologic malignancy is CML. In exemplaryembodiments, the cancer or hematologic malignancy is FL. In exemplaryembodiments, the cancer or hematologic malignancy is transformed mycosisfungoides. In exemplary embodiments, the cancer or hematologicmalignancy is Sézary syndrome. In exemplary embodiments, the cancer orhematologic malignancy is acute T-cell leukemia. In exemplaryembodiments, the cancer or hematologic malignancy is acute B-cellleukemia. In exemplary embodiments, the cancer or hematologic malignancyis Burkitt lymphoma. In exemplary embodiments, the cancer or hematologicmalignancy is myeloproliferative neoplasms. In exemplary embodiments,the cancer or hematologic malignancy is splenic marginal zone. Inexemplary embodiments, the cancer or hematologic malignancy is nodalmarginal zone. In exemplary embodiments, the cancer or hematologicmalignancy is extranodal marginal zone.

In one embodiment, the cancer or hematologic malignancy is a B celllymphoma. In a specific embodiment, provided herein is a method oftreating or managing a B cell lymphoma comprising administering to apatient a therapeutically effective amount of a compound providedherein, or a pharmaceutically acceptable derivative (e.g., salt orsolvate) thereof. Also provided herein is a method of treating orlessening one or more of the symptoms associated with a B cell lymphomacomprising administering to a patient a therapeutically effective amountof a compound provided herein, or a pharmaceutically acceptablederivative (e.g., salt or solvate) thereof. In one embodiment, the Bcell lymphoma is iNHL. In another embodiment, the B cell lymphoma isfollicular lymphoma. In another embodiment, the B cell lymphoma isWaldenstrom macroglobulinemia (lymphoplasmacytic lymphoma). In anotherembodiment, the B cell lymphoma is marginal zone lymphoma (MZL). Inanother embodiment, the B cell lymphoma is MCL. In another embodiment,the B cell lymphoma is HL. In another embodiment, the B cell lymphoma isaNHL. In another embodiment, the B cell lymphoma is DLBCL. In anotherembodiment, the B cell lymphoma is Richters lymphoma.

In one embodiment, the cancer or hematologic malignancy is a T celllymphoma. In a specific embodiment, provided herein is a method oftreating or managing a T cell lymphoma comprising administering to apatient a therapeutically effective amount of a compound providedherein, or a pharmaceutically acceptable derivative (e.g., salt orsolvate) thereof. Also provided herein is a method of treating orlessening one or more of the symptoms associated with a T cell lymphomacomprising administering to a patient a therapeutically effective amountof a compound provided herein, or a pharmaceutically acceptablederivative (e.g., salt or solvate) thereof. In one embodiment, the Tcell lymphoma is peripheral T cell lymphoma (PTCL). In anotherembodiment, the T cell lymphoma is cutaneous T cell lymphoma (CTCL).

In one embodiment, the cancer or hematologic malignancy is Sézarysyndrome. In a specific embodiment, provided herein is a method oftreating or managing Sézary syndrome comprising administering to apatient a therapeutically effective amount of a compound providedherein, or a pharmaceutically acceptable derivative (e.g., salt orsolvate) thereof. Also provided herein is a method of treating orlessening one or more of the symptoms associated with Sézary syndromecomprising administering to a patient a therapeutically effective amountof a compound provided herein, or a pharmaceutically acceptablederivative (e.g., salt or solvate) thereof. The symptoms associated withSézary syndrome include, but are not limited to, epidermotropism byneoplastic CD4+ lymphocytes, Pautrier's microabscesses, erythroderma,lymphadenopathy, atypical T cells in the peripheral blood, andhepatosplenomegalyIn one embodiment, the therapeutically effectiveamount for treating or managing Sézary syndrome is from about 25 mg to75 mg, administered twice daily. In other embodiments, thetherapeutically effective amount is from about 50 mg to about 75 mg,from about 30 mg to about 65 mg, from about 45 mg to about 60 mg, fromabout 30 mg to about 50 mg, or from about 55 mg to about 65 mg, each ofwhich is administered twice daily. In one embodiment, the effectiveamount is about 60 mg, administered twice daily.

In one embodiment, the cancer or hematologic malignancy is relapsed. Inone embodiment, the cancer or hematologic malignancy is refractory. Incertain embodiments, the cancer being treated or prevented is a specificsub-type of cancer described herein. In certain embodiments, thehematologic malignancy being treated or prevented is a specific sub-typeof hematologic malignancy described herein. Certain classifications oftype or sub-type of a cancer or hematologic malignancy provided hereinis known in the art. Without being limited by a particular theory, it isbelieved that many of the cancers that become relapsed or refractorydevelop resistance to the particular prior therapy administered to treatthe cancers. Thus, without being limited by a particular theory, acompound provided herein can provide a second line therapy by providingan alternative mechanism to treat cancers different from thosemechanisms utilized by certain prior therapies. Accordingly, in oneembodiment, provided herein is a method of treating or managing canceror hematologic malignancy comprising administering to a patient atherapeutically effective amount of a compound provided herein, or apharmaceutically acceptable derivative (e.g., salt or solvate) thereof,wherein the cancer or hematologic malignancy is relapsed after, orrefractory to, a prior therapy.

In exemplary embodiments, the cancer or hematologic malignancy isrefractory iNHL. In exemplary embodiments, the cancer or hematologicmalignancy is refractory CLL. In exemplary embodiments, the cancer orhematologic malignancy is refractory SLL. In exemplary embodiments, thecancer or hematologic malignancy is refractory to rituximab therapy. Inexemplary embodiments, the cancer or hematologic malignancy isrefractory to chemotherapy. In exemplary embodiments, the cancer orhematologic malignancy is refractory to radioimmunotherapy (RIT). Inexemplary embodiments, the cancer or hematologic malignancy is iNHL, FL,splenic marginal zone, nodal marginal zone, extranodal marginal zone, orSLL, the cancer or hematologic malignancy is refractory to rituximabtherapy, chemotherapy, and/or RIT.

In another exemplary embodiment, the cancer or hematologic malignancy islymphoma, and the cancer is relapsed after, or refractory to, thetreatment by a BTK inhibitor such as, but not limited to, ibrutinib orONO-4059. In another exemplary embodiment, the cancer or hematologicmalignancy is CLL, and the cancer is relapsed after, or refractory to,the treatment by a BTK inhibitor such as, but not limited to, ibrutiniband AVL-292.

In certain embodiments, provided herein are methods of treating orpreventing a solid tumor in a subject, comprising administering to thesubject a therapeutically effective amount of a compound provided herein(e.g., a compound of any of Formulae (I″), (I′), (A′), (I), (A), (II),(III), (IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII), (XIV),(XV), (XVI), and (XVII), e.g., a selective PI3K-γ inhibitor, e.g.,Compound 4), or a pharmaceutically acceptable form thereof.

In one embodiment, the solid tumor is selected from one or more of: acancer of the pulmonary system, a brain cancer, a cancer of thegastrointestinal tract, a skin cancer, a genitourinary cancer, apancreatic cancer, a lung cancer, a medullobastoma, a basal cellcarcinoma, a glioma, a breast cancer, a prostate cancer, a testicularcancer, an esophageal cancer, a hepatocellular cancer, a gastric cancer,a gastrointestinal stromal tumor (GIST), a colon cancer, a colorectalcancer, an ovarian cancer, a melanoma, a neuroectodermal tumor, head andneck cancer, a sarcoma, a soft-tissue sarcoma, fibrosarcoma,myxosarcoma, liposarcoma, a chondrosarcoma, an osteogenic sarcoma, achordoma, an angiosarcoma, an endotheliosarcoma, a lymphangiosarcoma, alymphangioendotheliosarcoma, a synovioma, a mesothelioma, aleiomyosarcoma, a cervical cancer, a uterine cancer, an endometrialcancer, a carcinoma, a bladder carcinoma, an epithelial carcinoma, asquamous cell carcinoma, an adenocarcinoma, a bronchogenic carcinoma, arenal cell carcinoma, a hepatoma, a bile duct carcinoma, aneuroendocrine cancer, a carcinoid tumor, diffuse type giant cell tumor,and glioblastoma.

In one embodiment, the compound, or a pharmaceutically acceptable formthereof, is administered at a dose sufficient to cause a decrease intumor growth of at least 10%, 20%, 30%, 40%, or 50% compared to areference value, after administration of the compound.

In one embodiment, the method further comprises administering animmunomodulator to the subject. In one embodiment, the immunomodulatoris a PDL-1 inhibitor or an anti-PDL-1 antibody.

In one embodiment, the method further comprises administering aPI3K-delta inhibitor to the subject.

In one embodiment, the compound, or a pharmaceutically acceptable formthereof, is administered at a dose such that the level of the compoundin the subject is higher than the compound's IC50 of PI3K-gammainhibition during at least 70%, 80%, 90%, 95%, 97%, 98%, or 99% of aselected time period, e.g., 6 hours, 12 hours, 24 hours, or 48 hours,immediately following the administration. In one embodiment, thecompound, or a pharmaceutically acceptable form thereof, is administeredat a dose such that the level of the compound in the subject is lowerthan the compound's IC50 of PI3K-delta inhibition during at least 70%,80%, 90%, 95%, 97%, 98%, or 99% of a selected time period, e.g., 6hours, 12 hours, 24 hours, or 48 hours, immediately following theadministration. In one embodiment, the level of the compound is measuredfrom the subject's plasma. In one embodiment, the level of the compoundis measured from the subject's tissue.

In one embodiment, the subject has been previously treated withcyclophosphamide, docetaxel, paclitaxel, 5-FU, or temozolomide.

In one embodiment, the anti-tumor effect of the compound is maintainedfor a period of time after the discontinuation of treatment with thecompound. In one embodiment, the period of time is at least 1 day, 2days, 3 days, 4 days, 5 days, or 6 days.

Treatment of an Inflammatory Disorder

In one embodiment, provided herein is a method of treating aninflammation disorder, including autoimmune diseases in a subject. Themethod comprises administering to said subject a therapeuticallyeffective amount of a compound provided herein, or a pharmaceuticallyacceptable form (e.g., pharmaceutically acceptable salts, hydrates,solvates, isomers, prodrugs, and isotopically labeled derivatives)thereof, or a pharmaceutical composition as provided herein. Examples ofautoimmune diseases include but are not limited to acute disseminatedencephalomyelitis (ADEM), Addison's disease, antiphospholipid antibodysyndrome (APS), aplastic anemia, autoimmune hepatitis, autoimmune skindisease, coeliac disease, Crohn's disease, Diabetes mellitus (type 1),Goodpasture's syndrome, Graves' disease, Guillain-Barré syndrome (GBS),Hashimoto's disease, lupus erythematosus, multiple sclerosis, myastheniagravis, opsoclonus myoclonus syndrome (OMS), optic neuritis, Ord'sthyroiditis, oemphigus, polyarthritis, primary biliary cirrhosis,psoriasis, rheumatoid arthritis, Reiter's syndrome, Takayasu'sarteritis, temporal arteritis (also known as “giant cell arteritis”),warm autoimmune hemolytic anemia, Wegener's granulomatosis, alopeciauniversalis (e.g., inflammatory alopecia), Chagas disease, chronicfatigue syndrome, dysautonomia, endometriosis, hidradenitis suppurativa,interstitial cystitis, neuromyotonia, sarcoidosis, scleroderma,ulcerative colitis, vitiligo, and vulvodynia. Other disorders includebone-resorption disorders and thrombosis.

Inflammation takes on many forms and includes, but is not limited to,acute, adhesive, atrophic, catarrhal, chronic, cirrhotic, diffuse,disseminated, exudative, fibrinous, fibrosing, focal, granulomatous,hyperplastic, hypertrophic, interstitial, metastatic, necrotic,obliterative, parenchymatous, plastic, productive, proliferous,pseudomembranous, purulent, sclerosing, seroplastic, serous, simple,specific, subacute, suppurative, toxic, traumatic, and/or ulcerativeinflammation.

Exemplary inflammatory conditions include, but are not limited to,inflammation associated with acne, anemia (e.g., aplastic anemia,haemolytic autoimmune anaemia), asthma, arteritis (e.g., polyarteritis,temporal arteritis, periarteritis nodosa, Takayasu's arteritis),arthritis (e.g., crystalline arthritis, osteoarthritis, psoriaticarthritis, gout flare, gouty arthritis, reactive arthritis, rheumatoidarthritis and Reiter's arthritis), ankylosing spondylitis, amylosis,amyotrophic lateral sclerosis, autoimmune diseases, allergies orallergic reactions, atherosclerosis, bronchitis, bursitis, chronicprostatitis, conjunctivitis, Chagas disease, chronic obstructivepulmonary disease, cermatomyositis, diverticulitis, diabetes (e.g., typeI diabetes mellitus, type 2 diabetes mellitus), a skin condition (e.g.,psoriasis, eczema, burns, dermatitis, pruritus (itch)), endometriosis,Guillain-Barre syndrome, infection, ischaemic heart disease, Kawasakidisease, glomerulonephritis, gingivitis, hypersensitivity, headaches(e.g., migraine headaches, tension headaches), ileus (e.g.,postoperative ileus and ileus during sepsis), idiopathicthrombocytopenic purpura, interstitial cystitis (painful bladdersyndrome), gastrointestinal disorder (e.g., selected from peptic ulcers,regional enteritis, diverticulitis, gastrointestinal bleeding,eosinophilic gastrointestinal disorders (e.g., eosinophilic esophagitis,eosinophilic gastritis, eosinophilic gastroenteritis, eosinophiliccolitis), gastritis, diarrhea, gastroesophageal reflux disease (GORD, orits synonym GERD), inflammatory bowel disease (IBD) (e.g., Crohn'sdisease, ulcerative colitis, collagenous colitis, lymphocytic colitis,ischaemic colitis, diversion colitis, Behcet's syndrome, indeterminatecolitis) and inflammatory bowel syndrome (IBS)), lupus, multiplesclerosis, morphea, myeasthenia gravis, myocardial ischemia, nephroticsyndrome, pemphigus vulgaris, pernicious aneaemia, peptic ulcers,polymyositis, primary biliary cirrhosis, neuroinflammation associatedwith brain disorders (e.g., Parkinson's disease, Huntington's disease,and Alzheimer's disease), prostatitis, chronic inflammation associatedwith cranial radiation injury, pelvic inflammatory disease, polymyalgiarheumatic, reperfusion injury, regional enteritis, rheumatic fever,systemic lupus erythematosus, scleroderma, scierodoma, sarcoidosis,spondyloarthopathies, Sjogren's syndrome, thyroiditis, transplantationrejection, tendonitis, trauma or injury (e.g., frostbite, chemicalirritants, toxins, scarring, burns, physical injury), vasculitis,vitiligo and Wegener's granulomatosis. In certain embodiments, theinflammatory disorder is selected from arthritis (e.g., rheumatoidarthritis), inflammatory bowel disease, inflammatory bowel syndrome,asthma, psoriasis, endometriosis, interstitial cystitis andprostatistis. In certain embodiments, the inflammatory condition is anacute inflammatory condition (e.g., for example, inflammation resultingfrom infection). In certain embodiments, the inflammatory condition is achronic inflammatory condition (e.g., conditions resulting from asthma,arthritis and inflammatory bowel disease). The compounds can also beuseful in treating inflammation associated with trauma andnon-inflammatory myalgia.

Immune disorders, such as auto-immune disorders, include, but are notlimited to, arthritis (including rheumatoid arthritis,spondyloarthopathies, gouty arthritis, degenerative joint diseases suchas osteoarthritis, systemic lupus erythematosus, Sjogren's syndrome,ankylosing spondylitis, undifferentiated spondylitis, Behcet's disease,haemolytic autoimmune anaemias, multiple sclerosis, amyotrophic lateralsclerosis, amylosis, acute painful shoulder, psoriatic, and juvenilearthritis), asthma, atherosclerosis, osteoporosis, bronchitis,tendonitis, bursitis, skin condition (e.g., psoriasis, eczema, burns,dermatitis, pruritus (itch)), enuresis, eosinophilic disease,gastrointestinal disorder (e.g., selected from peptic ulcers, regionalenteritis, diverticulitis, gastrointestinal bleeding, eosinophilicgastrointestinal disorders (e.g., eosinophilic esophagitis, eosinophilicgastritis, eosinophilic gastroenteritis, eosinophilic colitis),gastritis, diarrhea, gastroesophageal reflux disease (GORD, or itssynonym GERD), inflammatory bowel disease (IBD) (e.g., Crohn's disease,ulcerative colitis, collagenous colitis, lymphocytic colitis, ischaemiccolitis, diversion colitis, Behcet's syndrome, indeterminate colitis)and inflammatory bowel syndrome (IBS)), relapsing polychondritis (e.g.,atrophic polychondritis and systemic polychondromalacia), and disordersameliorated by a gastroprokinetic agent (e.g., ileus, postoperativeileus and ileus during sepsis; gastroesophageal reflux disease (GORD, orits synonym GERD); eosinophilic esophagitis, gastroparesis such asdiabetic gastroparesis; food intolerances and food allergies and otherfunctional bowel disorders, such as non-ulcerative dyspepsia (NUD) andnon-cardiac chest pain (NCCP, including costo-chondritis)). In certainembodiments, a method of treating inflammatory or autoimmune diseases isprovided comprising administering to a subject (e.g., a mammal) atherapeutically effective amount of a compound provided herein, or apharmaceutically acceptable form (e.g., pharmaceutically acceptablesalts, hydrates, solvates, isomers, prodrugs, and isotopically labeledderivatives) thereof, or a pharmaceutical composition as providedherein, that selectively inhibit PI3K-δ and/or PI3K-γ as compared to allother type I PI3 kinases. Such selective inhibition of PI3K-δ and/orPI3K-γ can be advantageous for treating any of the diseases orconditions described herein. For example, selective inhibition of PI3K-δand/or PI3K-γ can inhibit inflammatory responses associated withinflammatory diseases, autoimmune disease, or diseases related to anundesirable immune response including, but not limited to asthma,emphysema, allergy, dermatitis, rheumatoid arthritis, psoriasis, lupuserythematosus, anaphylaxsis, or graft versus host disease. Selectiveinhibition of PI3K-δ and/or PI3K-γ can further provide for a reductionin the inflammatory or undesirable immune response without a concomitantreduction in the ability to reduce a bacterial, viral, and/or fungalinfection. Selective inhibition of both PI3K-δ and PI3K-γ can beadvantageous for inhibiting the inflammatory response in the subject toa greater degree than that would be provided for by inhibitors thatselectively inhibit PI3K-δ or PI3K-γ alone. In one aspect, one or moreof the subject methods are effective in reducing antigen specificantibody production in vivo by about 2-fold, 3-fold, 4-fold, 5-fold,7.5-fold, 10-fold, 25-fold, 50-fold, 100-fold, 250-fold, 500-fold,750-fold, or about 1000-fold or more. In another aspect, one or more ofthe subject methods are effective in reducing antigen specific IgG3and/or IgGM production in vivo by about 2-fold, 3-fold, 4-fold, 5-fold,7.5-fold, 10-fold, 25-fold, 50-fold, 100-fold, 250-fold, 500-fold,750-fold, or about 1000-fold or more.

In one aspect, one of more of the subject methods are effective inameliorating symptoms associated with rheumatoid arthritis including,but not limited to a reduction in the swelling of joints, a reduction inserum anti-collagen levels, and/or a reduction in joint pathology suchas bone resorption, cartilage damage, pannus, and/or inflammation. Inanother aspect, the subject methods are effective in reducing ankleinflammation by at least about 2%, 5%, 10%, 15%, 20%, 25%, 30%, 50%, or60%, or about 75% to 90%. In another aspect, the subject methods areeffective in reducing knee inflammation by at least about 2%, 5%, 10%,15%, 20%, 25%, 30%, 50%, or 60%, or about 75% to 90% or more. In stillanother aspect, the subject methods are effective in reducing serumanti-type II collagen levels by at least about 10%, 12%, 15%, 20%, 24%,25%, 30%, 35%, 50%, 60%, 75%, 80%, 86%, or 87%, or about 90% or more. Inanother aspect, the subject methods are effective in reducing anklehistopathology scores by about 5%, 10%, 15%, 20%, 25%, 30%, 40%, 50%,60%, 75%, 80%, or 90%, or more. In still another aspect, the subjectmethods are effective in reducing knee histopathology scores by about5%, 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60%, 75%, 80%, or 90%, or more.

In certain embodiments, provided herein are methods of treating orpreventing arthritis in a subject, comprising administering to thesubject a therapeutically effective amount of a compound provided herein(e.g., a compound of any of Formulae (I″), (I′), (A′), (I), (A), (II),(III), (IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII), (XIV),(XV), (XVI), and (XVII), e.g., a selective PI3K-γ inhibitor, e.g.,Compound 4), or a pharmaceutically acceptable form thereof. In oneembodiment, the treatment results in reduction of periosteal boneformation in the subject. In one embodiment, the treatment results in atleast 10%, 20%, 40%, 47%, 50%, 52%, 60%, 80%, or 82% reduction ofperiosteal bone formation in the subject, compared to a reference value.In one embodiment, the periosteal bone formation is measured byhistopathology score or periosteal bone width. In one embodiment, thetreatment results in at least 10%, 20%, 27%, 30%, 36%, 40%, 45%, 50%, or57% reduction of inflammation, at least 10%, 20%, 28%, 30%, 40%, 44%,50%, or 60%, 70%, or 71% reduction of pannus, at least 10%, 20%, 28%,30%, 40%, 45%, 50%, or 59% reduction of cartilage damage, or at least10%, 20%, 25%, 30%, 40%, 44%, 50%, 60%, or 65% reduction of boneresorption in the subject, compared to a reference value. In oneembodiment, wherein the treatment results in reduction of joint swellingor anti-collagen level in the subject.

In some embodiments, provided herein are methods for treating disordersor conditions in which the δ isoform of PI3K is implicated to a greaterextent than other PI3K isoforms such as PI3K-α and/or PI3K-β. In someembodiments, provided herein are methods for treating disorders orconditions in which the γ isoform of PI3K is implicated to a greaterextent than other PI3K isoforms such as PI3K-α and/or PI3K-β. Selectiveinhibition of PI3K-δ and/or PI3K-γ can provide advantages over usingless selective compounds which inhibit PI3K-α and/or PI3K-β, such as animproved side effects profile or lessened reduction in the ability toreduce a bacterial, viral, and/or fungal infection.

In other embodiments, provided herein are methods of using a compoundprovided herein, or a pharmaceutically acceptable form (e.g.,pharmaceutically acceptable salts, hydrates, solvates, isomers,prodrugs, and isotopically labeled derivatives) thereof, or apharmaceutical composition as provided herein, to treat respiratorydiseases including, but not limited to, diseases affecting the lobes oflung, pleural cavity, bronchial tubes, trachea, upper respiratory tract,or the nerves and muscle for breathing. For example, methods areprovided to treat obstructive pulmonary disease. Chronic obstructivepulmonary disease (COPD) is an umbrella term for a group of respiratorytract diseases that are characterized by airflow obstruction orlimitation. Conditions included in this umbrella term include, but arenot limited to: chronic bronchitis, emphysema, and bronchiectasis.

In another embodiment, a compound provided herein, or a pharmaceuticallyacceptable form (e.g., pharmaceutically acceptable salts, hydrates,solvates, isomers, prodrugs, and isotopically labeled derivatives)thereof, or a pharmaceutical composition as provided herein is used forthe treatment of asthma. Also, a compound provided herein, or apharmaceutically acceptable form thereof, or a pharmaceuticalcomposition described herein, can be used for the treatment ofendotoxemia and sepsis. In one embodiment, the compounds orpharmaceutical compositions described herein are used to for thetreatment of rheumatoid arthritis (RA). In yet another embodiment, thecompounds or pharmaceutical compositions described herein is used forthe treatment of contact or atopic dermatitis. Contact dermatitisincludes irritant dermatitis, phototoxic dermatitis, allergicdermatitis, photoallergic dermatitis, contact urticaria, systemiccontact-type dermatitis and the like. Irritant dermatitis can occur whentoo much of a substance is used on the skin of when the skin issensitive to certain substance. Atopic dermatitis, sometimes calledeczema, is a kind of dermatitis, an atopic skin disease.

In certain embodiments, provided herein are methods of reducingneutrophil migration or infiltration in a subject suffering from aninflammatory disease, comprising administering to the subject atherapeutically effective amount of a compound provided herein (e.g., acompound of any of Formulae (I″), (I′), (A′), (I), (A), (II), (III),(IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII), (XIV), (XV),(XVI), and (XVII), e.g., a selective PI3K-γ inhibitor, e.g., Compound4), or a pharmaceutically acceptable form thereof. In one embodiment,the neutrophil migration or infiltration is reduced by at least about10%, 20%, 40%, 60%, 80%, or 90% compared to a reference value, afteradministration of the compound. In one embodiment, the inflammatorydisease is selected from the group consisting of COPD, arthritis,asthma, psoriasis, scleroderma, myositis, sarcoidosis, dermatomyositis,CREST syndrome, systemic lupus erythematosus, Sjorgren syndrome,encephalomyelitis, and inflammatory bowel disease (IBD). In oneembodiment, the inflammatory disease is COPD or arthritis. In oneembodiment, the subject is unresponsive or refractory to a PI3K-deltainhibitor treatment.

Treatment of Other Disorders or Conditions

In some embodiments, the disclosure provides a method of treatingdiseases related to vasculogenesis or angiogenesis in a subject thatcomprises administering to said subject a therapeutically effectiveamount of a compound provided herein, or a pharmaceutically acceptableform (e.g., pharmaceutically acceptable salts, hydrates, solvates,isomers, prodrugs, and isotopically labeled derivatives) thereof, or apharmaceutical composition as provided herein. In some embodiments, saidmethod is for treating a disease selected from tumor angiogenesis,chronic inflammatory disease such as rheumatoid arthritis and chronicinflammatory demyelinating polyneuropathy, atherosclerosis, inflammatorybowel disease, skin diseases such as psoriasis, eczema, and scleroderma,diabetes, diabetic retinopathy, retinopathy of prematurity, age-relatedmacular degeneration, hemangioma, glioma, melanoma, Kaposi's sarcoma andovarian, breast, lung, pancreatic, prostate, colon and epidermoidcancer.

In addition, the compounds described herein can be used for thetreatment of arteriosclerosis, including atherosclerosis.Arteriosclerosis is a general term describing any hardening of medium orlarge arteries. Atherosclerosis is a hardening of an artery specificallydue to an atheromatous plaque.

In some embodiments, provided herein is a method of treating acardiovascular disease in a subject that comprises administering to saidsubject a therapeutically effective amount of a compound providedherein, or a pharmaceutically acceptable form (e.g., pharmaceuticallyacceptable salts, hydrates, solvates, isomers, prodrugs, andisotopically labeled derivatives) thereof, or a pharmaceuticalcomposition as provided herein. Examples of cardiovascular conditionsinclude, but are not limited to, atherosclerosis, restenosis, vascularocclusion and carotid obstructive disease.

In some embodiments, the disclosure relates to a method of treatingdiabetes in a subject that comprises administering to said subject atherapeutically effective amount of a compound provided herein, or apharmaceutically acceptable form (e.g., pharmaceutically acceptablesalts, hydrates, solvates, isomers, prodrugs, and isotopically labeledderivatives) thereof, or a pharmaceutical composition as providedherein.

In addition, a compound provided herein, or a pharmaceuticallyacceptable form (e.g., pharmaceutically acceptable salts, hydrates,solvates, isomers, prodrugs, and isotopically labeled derivatives)thereof, or a pharmaceutical composition as provided herein, can be usedto treat acne. In certain embodiments, the inflammatory condition and/orimmune disorder is a skin condition. In some embodiments, the skincondition is pruritus (itch), psoriasis, eczema, burns or dermatitis. Incertain embodiments, the skin condition is psoriasis. In certainembodiments, the skin condition is pruritis.

In certain embodiments, the inflammatory disorder and/or the immunedisorder is a gastrointestinal disorder. In some embodiments, thegastrointestinal disorder is selected from gastrointestinal disorder(e.g., selected from peptic ulcers, regional enteritis, diverticulitis,gastrointestinal bleeding, eosinophilic gastrointestinal disorders(e.g., eosinophilic esophagitis, eosinophilic gastritis, eosinophilicgastroenteritis, eosinophilic colitis), gastritis, diarrhea,gastroesophageal reflux disease (GORD, or its synonym GERD),inflammatory bowel disease (IBD) (e.g., Crohn's disease, ulcerativecolitis, collagenous colitis, lymphocytic colitis, ischaemic colitis,diversion colitis, Behcet's syndrome, indeterminate colitis) andinflammatory bowel syndrome (IBS)). In certain embodiments, thegastrointestinal disorder is inflammatory bowel disease (IBD).

Further, a compound provided herein, or a pharmaceutically acceptableform (e.g., pharmaceutically acceptable salts, hydrates, solvates,isomers, prodrugs, and isotopically labeled derivatives) thereof, or apharmaceutical composition as provided herein, can be used for thetreatment of glomerulonephritis. Glomerulonephritis is a primary orsecondary autoimmune renal disease characterized by inflammation of theglomeruli. It can be asymptomatic, or present with hematuria and/orproteinuria. There are many recognized types, divided in acute, subacuteor chronic glomerulonephritis. Causes are infectious (bacterial, viralor parasitic pathogens), autoimmune or paraneoplastic.

In some embodiments, provided herein are compounds, or pharmaceuticallyacceptable forms (e.g., pharmaceutically acceptable salts, hydrates,solvates, isomers, prodrugs, and isotopically labeled derivatives)thereof, or pharmaceutical compositions as provided herein, for thetreatment of multiorgan failure. Also provided herein are compounds, orpharmaceutically acceptable forms (e.g., pharmaceutically acceptablesalts, hydrates, solvates, isomers, prodrugs, and isotopically labeledderivatives) thereof, or pharmaceutical compositions as provided herein,for the treatment of liver diseases (including diabetes), gall bladderdisease (including gallstones), pancreatitis or kidney disease(including proliferative glomerulonephritis and diabetes-induced renaldisease) or pain in a subject.

In some embodiments, provided herein are compounds, or pharmaceuticallyacceptable forms (e.g., pharmaceutically acceptable salts, hydrates,solvates, isomers, prodrugs, and isotopically labeled derivatives)thereof, or pharmaceutical compositions as provided herein, for theprevention of blastocyte implantation in a subject.

In some embodiments, provided herein are compounds, or pharmaceuticallyacceptable forms (e.g., pharmaceutically acceptable salts, hydrates,solvates, isomers, prodrugs, and isotopically labeled derivatives)thereof, or pharmaceutical compositions as provided herein, for thetreatment of disorders involving platelet aggregation or plateletadhesion, including, but not limited to, Idiopathic thrombocytopenicpurpura, Bernard-Soulier syndrome, Glanzmann's thrombasthenia, Scott'ssyndrome, von Willebrand disease, Hermansky-Pudlak Syndrome, and Grayplatelet syndrome.

In some embodiments, provided herein are compounds, or pharmaceuticallyacceptable forms (e.g., pharmaceutically acceptable salts, hydrates,solvates, isomers, prodrugs, and isotopically labeled derivatives)thereof, or pharmaceutical compositions as provided herein, for thetreatment of a disease which is skeletal muscle atrophy, skeletal ormuscle hypertrophy. In some embodiments, provided herein are compounds,or pharmaceutically acceptable forms (e.g., pharmaceutically acceptablesalts, hydrates, solvates, isomers, prodrugs, and isotopically labeledderivatives) thereof, or pharmaceutical compositions as provided herein,for the treatment of disorders that include, but are not limited to,cancers as discussed herein, transplantation-related disorders (e.g.,lowering rejection rates, graft-versus-host disease, etc.), muscularsclerosis (MS), allergic disorders (e.g., arthritis, allergicencephalomyelitis) and other immunosuppressive-related disorders,metabolic disorders (e.g., diabetes), reducing intimal thickeningfollowing vascular injury, and misfolded protein disorders (e.g.,Alzheimer's Disease, Gaucher's Disease, Parkinson's Disease,Huntington's Disease, cystic fibrosis, macular degeneration, retinitispigmentosa, and prion disorders) (as mTOR inhibition can alleviate theeffects of misfolded protein aggregates). The disorders also includehamartoma syndromes, such as tuberous sclerosis and Cowden Disease (alsotermed Cowden syndrome and multiple hamartoma syndrome).

Additionally, a compound provided herein, or a pharmaceuticallyacceptable form (e.g., pharmaceutically acceptable salts, hydrates,solvates, isomers, prodrugs, and isotopically labeled derivatives)thereof, or a pharmaceutical composition as provided herein, can be usedfor the treatment of bursitis, lupus, acute disseminatedencephalomyelitis (ADEM), Addison's disease, antiphospholipid antibodysyndrome (APS), amyloidosis (including systemic and localizedamyloidosis; and primary and secondary amyloidosis), aplastic anemia,autoimmune hepatitis, coeliac disease, crohn's disease, diabetesmellitus (type 1), eosinophilic gastroenterides, goodpasture's syndrome,graves' disease, guillain-barré syndrome (GBS), hashimoto's disease,inflammatory bowel disease, lupus erythematosus (including cutaneouslupus erythematosus and systemic lupus erythematosus), myastheniagravis, opsoclonus myoclonus syndrome (OMS), optic neuritis, ord'sthyroiditis, ostheoarthritis, uveoretinitis, pemphigus, polyarthritis,primary biliary cirrhosis, reiter's syndrome, takayasu's arteritis,temporal arteritis, warm autoimmune hemolytic anemia, wegener'sgranulomatosis, alopecia universalis, chagas' disease, chronic fatiguesyndrome, dysautonomia, endometriosis, hidradenitis suppurativa,interstitial cystitis, neuromyotonia, sarcoidosis, scleroderma,ulcerative colitis, vitiligo, vulvodynia, appendicitis, arteritis,arthritis, blepharitis, bronchiolitis, bronchitis, cervicitis,cholangitis, cholecystitis, chorioamnionitis, colitis, conjunctivitis,cystitis, dacryoadenitis, dermatomyositis, endocarditis, endometritis,enteritis, enterocolitis, epicondylitis, epididymitis, fasciitis,fibrositis, gastritis, gastroenteritis, gingivitis, hepatitis,hidradenitis, ileitis, iritis, laryngitis, mastitis, meningitis,myelitis, myocarditis, myositis, nephritis, omphalitis, oophoritis,orchitis, osteitis, otitis, pancreatitis, parotitis, pericarditis,peritonitis, pharyngitis, pleuritis, phlebitis, pneumonitis, proctitis,prostatitis, pyelonephritis, rhinitis, salpingitis, sinusitis,stomatitis, synovitis, pigmented villonodular synovitis (also known astenosynovial giant cell tumor), tendonitis, tonsillitis, uveitis (e.g.,ocular uveitis), vaginitis, vasculitis, or vulvitis.

Further, the compounds provided herein may be used for the treatment ofPerennial allergic rhinitis, Mesenteritis, Peritonitis, Acrodermatitis,Angiodermatitis, Atopic dermatitis, Contact dermatitis, Eczema, Erythemamultiforme, Intertrigo, Stevens Johnson syndrome, Toxic epidermalnecrolysis, Skin allergy, Severe allergic reaction/anaphylaxis, Allergicgranulomatosis, Wegener granulomatosis, Allergic conjunctivitis,Chorioretinitis, Conjunctivitis, Infectious keratoconjunctivitis,Keratoconjunctivitis, Ophthalmia neonatorum, Trachoma, Uveitis, Ocularinflammation, ocular lymphoma, MALT lymphoma, Blepharoconjunctivitis,Mastitis, Gingivitis, Pericoronitis, Pharyngitis, Rhinopharyngitis,Sialadenitis, Musculoskeletal system inflammation, Adult onset Stillsdisease, Behcets disease, Bursitis, Chondrocalcinosis, Dactylitis, Feltysyndrome, Gout, Infectious arthritis, Lyme disease, Inflammatoryosteoarthritis, Periarthritis, Reiter syndrome, Ross River virusinfection, Acute Respiratory, Distress Syndrome, Acute bronchitis, Acutesinusitis, Allergic rhinitis, Asthma, Severe refractory asthma,Pharyngitis, Pleurisy, Rhinopharyngitis, Seasonal allergic rhinitis,Sinusitis, Status asthmaticus, Tracheobronchitis, Rhinitis, Serositis,Meningitis, Neuromyelitis optica, Poliovirus infection, Alport syndrome,Balanitis, Epididymitis, Epididymo orchitis, Focal segmental,Glomerulosclerosis, Glomerulonephritis, IgA Nephropathy (Berger'sDisease), Orchitis, Parametritis, Pelvic inflammatory disease,Prostatitis, Pyelitis, Pyelocystitis, Pyelonephritis, Wegenergranulomatosis, Hyperuricemia, Aortitis, Arteritis, Chylopericarditis,Dressler syndrome, Endarteritis, Endocarditis, Extracranial temporalarteritis, HIV associated arteritis, Intracranial temporal arteritis,Kawasaki disease, Lymphangiophlebitis, Mondor disease, Periarteritis, orPericarditis.

In other aspects, the compounds provided herein are used for thetreatment of Autoimmune hepatitis, Jejunitis, Mesenteritis, Mucositis,Non alcoholic steatohepatitis, Non viral hepatitis, Autoimmunepancreatitis, Perihepatitis, Peritonitis, Pouchitis, Proctitis,Pseudomembranous colitis, Rectosigmoiditis, Salpingoperitonitis,Sigmoiditis, Steatohepatitis, Ulcerative colitis, Churg Strausssyndrome, Ulcerative proctitis, Irritable bowel syndrome,Gastrointestinal inflammation, Acute enterocolitis, Anusitis, Balsernecrosis, Cholecystitis, Colitis, Crohns disease, Diverticulitis,Enteritis, Enterocolitis, Enterohepatitis, Eosinophilic esophagitis,Esophagitis, Gastritis, Hemorrhagic enteritis, Hepatitis, Hepatitisvirus infection, Hepatocholangitis, Hypertrophic gastritis, Ileitis,Ileocecitis, Sarcoidosis, Inflammatory bowel disease, Ankylosingspondylitis, Rheumatoid arthritis, Juvenile rheumatoid arthritis,Psoriasis, Psoriatic arthritis, Lupus (cutaneous/systemic/nephritis),AIDS, Agammaglobulinemia, AIDS related complex, Brutons disease, ChediakHigashi syndrome, Common variable immunodeficiency, DiGeorge syndrome,Dysgammaglobulinemia, Immunoglobulindeficiency, Job syndrome, Nezelofsyndrome, Phagocyte bactericidal disorder, Wiskott Aldrich syndrome,Asplenia, Elephantiasis, Hypersplenism, Kawasaki disease,Lymphadenopathy, Lymphedema, Lymphocele, Nonne Milroy Meige syndrome,Spleen disease, Splenomegaly, Thymoma, Thymus disease, Perivasculitis,Phlebitis, Pleuropericarditis, Polyarteritis nodosa, Vasculitis,Takayasus arteritis, Temporal arteritis, Thromboangiitis,Thromboangiitis obliterans, Thromboendocarditis, Thrombophlebitis, orCOPD.

In another aspect, provided herein are methods of disrupting thefunction of a leukocyte or disrupting a function of an osteoclast. Themethod includes contacting the leukocyte or the osteoclast with afunction disrupting amount of a compound provided herein.

In another aspect, provided herein are methods for the treatment of anophthalmic disease by administering one or more of compounds providedherein, or pharmaceutically acceptable forms thereof, or pharmaceuticalcompositions as provided herein, to the eye of a subject.

Methods are further provided for administering the compounds providedherein via eye drop, intraocular injection, intravitreal injection,topically, or through the use of a drug eluting device, microcapsule,implant, or microfluidic device. In some cases, the compounds providedherein are administered with a carrier or excipient that increases theintraocular penetrance of the compound such as an oil and water emulsionwith colloid particles having an oily core surrounded by an interfacialfilm.

In certain embodiments, provided herein are methods of treating,preventing, and/or managing a disease or a disorder using a compound, ora pharmaceutically acceptable form (e.g., pharmaceutically acceptablesalts, hydrates, solvates, isomers, prodrugs, and isotopically labeledderivatives) thereof, or pharmaceutical compositions as provided herein,wherein the disease or disorder is: Crohn's disease; cutaneous lupus;multiple sclerosis; rheumatoid arthritis; and systemic lupuserythematosus.

In other embodiments, provided herein are methods of treating,preventing and/or managing a disease or a disorder using a compound, ora pharmaceutically acceptable form (e.g., pharmaceutically acceptablesalts, hydrates, solvates, isomers, prodrugs, and isotopically labeledderivatives) thereof, or pharmaceutical compositions as provided herein,wherein the disease or disorder is: ankylosing spondylitis; chronicobstructive pulmonary disease; myasthenia gravis; ocular uveitis,psoriasis; and psoriatic arthritis.

In other embodiments, provided herein are methods of treating,preventing and/or managing a disease or a disorder using a compound, ora pharmaceutically acceptable form (e.g., pharmaceutically acceptablesalts, hydrates, solvates, isomers, prodrugs, and isotopically labeledderivatives) thereof, or pharmaceutical compositions as provided herein,wherein the disease or disorder is: adult-onset Still's disease;inflammatory alopecia; amyloidosis; antiphospholipid syndrome;autoimmune hepatitis; autoimmune skin disease, Behcet's disease; chronicinflammatory demyelinating polyneuropathy; eosinophilic gastroenteritis;inflammatory myopathies, pemphigus, polymyalgia rheumatica; relapsingpolychondritis; Sjorgen's syndrome; temporal arthritis; ulcerativecolitis; vasculis; vitiligo, and Wegner's granulomatosis.

In other embodiments, provided herein are methods of treating,preventing and/or managing a disease or a disorder using a compound, ora pharmaceutically acceptable form (e.g., pharmaceutically acceptablesalts, hydrates, solvates, isomers, prodrugs, and isotopically labeledderivatives) thereof, or pharmaceutical compositions as provided herein,wherein the disease or disorder is: gout flare; sacoidosis; and systemicsclerosis.

In certain embodiments, provided herein are methods of treating,preventing and/or managing a disease or a disorder using a compound, ora pharmaceutically acceptable form (e.g., pharmaceutically acceptablesalts, hydrates, solvates, isomers, prodrugs, and isotopically labeledderivatives) thereof, or pharmaceutical compositions as provided herein,wherein the disease or disorder is: asthma; arthritis (e.g., rheumatoidarthritis and psoriatic arthritis); psoriasis; scleroderma; myositis(e.g., dermatomyositis); lupus (e.g., cutaneous lupus erythematosus(“CLE”) or systemic lupus erythematosus (“SLE”)); or Sjögren's syndrome.

Efficacy of a compound provided herein in treating, preventing and/ormanaging the disease or disorder can be tested using various animalmodels known in the art. For example: efficacy in treating, preventingand/or managing asthma can be assessed using ova induced asthma modeldescribed, for example, in Lee et al. (2006) J Allergy Clin Immunol118(2):403-9; efficacy in treating, preventing and/or managing arthritis(e.g., rheumatoid or psoriatic arthritis) can be assessed usingautoimmune animal models described, for example, in Williams et al.(2010) Chem Biol, 17(2):123-34, WO 2009/088986, WO2009/088880, and WO2011/008302; efficacy in treating, preventing and/or managing psoriasiscan be assessed using transgenic or knockout mouse model with targetedmutations in epidermis, vasculature or immune cells, mouse modelresulting from spontaneous mutations, and immuno-deficient mouse modelwith xenotransplantation of human skin or immune cells, all of which aredescribed, for example, in Boehncke et al. (2007) Clinics inDermatology, 25: 596-605; efficacy in treating, preventing and/ormanaging fibrosis or fibrotic condition can be assessed using theunilateral ureteral obstruction model of renal fibrosis (see Chevalieret al., Kidney International (2009) 75:1145-1152), the bleomycin inducedmodel of pulmonary fibrosis (see Moore and Hogaboam, Am. J. Physiol.Lung. Cell. Mol. Physiol. (2008) 294:L152-L160), a variety ofliver/biliary fibrosis models (see Chuang et al., Clin Liver Dis (2008)12:333-347 and Omenetti, A. et al. (2007) Laboratory Investigation87:499-514 (biliary duct-ligated model)), or a number of myelofibrosismouse models (see Varicchio, L. et al. (2009) Expert Rev. Hematol.2(3):315-334); efficacy in treating, preventing and/or managingscleroderma can be assessed using mouse model induced by repeated localinjections of bleomycin (“BLM”) described, for example, in Yamamoto etal. (1999) J Invest Dermatol 112: 456-462; efficacy in treating,preventing and/or managing dermatomyositis can be assessed usingmyositis mouse model induced by immunization with rabbit myosindescribed, for example, in Phyanagi et al. (2009) Arthritis &Rheumatism, 60(10): 3118-3127; efficacy in treating, preventing and/ormanaging lupus (e.g., CLE or SLE) can be assessed using various animalmodels described, for example, in Ghoreishi et al. (2009) Lupus, 19:1029-1035, Ohl et al. (2011) Journal of Biomedicine and Biotechnology,Article ID 432595 (14 pages), Xia et al. (2011) Rheumatology,50:2187-2196, Pau et al. (2012) PLoS ONE, 7(5):e36761 (15 pages),Mustafa et al. (2011) Toxicology, 290:156-168, Ichikawa et al. (2012)Arthritis and Rheumatism, 62(2): 493-503, Ouyang et al. (2012) J MolMed, DOI 10.1007/s00109-012-0866-3 (10 pages), Rankin et al. (2012)Journal of Immunology, 188:1656-1667; and efficacy in treating,preventing and/or managing Sjögren's syndrome can be assessed usingvarious mouse models described, for example, in Chiorini et al. (2009)Journal of Autoimmunity, 33: 190-196.

In one embodiment, provided herein is a method of treating, preventingand/or managing asthma. As used herein, “asthma” encompasses airwayconstriction regardless of the cause. Common triggers of asthma include,but are not limited to, exposure to an environmental stimulants (e.g.,allergens), cold air, warm air, perfume, moist air, exercise orexertion, and emotional stress. Also provided herein is a method oftreating, preventing and/or managing one or more symptoms associatedwith asthma. Examples of the symptoms include, but are not limited to,severe coughing, airway constriction and mucus production.

In one embodiment, provided herein is a method of treating, preventingand/or managing arthritis. As used herein, “arthritis” encompasses alltypes and manifestations of arthritis. Examples include, but are notlimited to, crystalline arthritis, osteoarthritis, psoriatic arthritis,gouty arthritis, reactive arthritis, rheumatoid arthritis and Reiter'sarthritis. In one embodiment, the disease or disorder is rheumatoidarthritis. In another embodiment, the disease or disorder is psoriaticarthritis. Also provided herein is a method of treating, preventingand/or managing one or more symptoms associated with arthritis. Examplesof the symptoms include, but are not limited to, joint pain, whichprogresses into joint deformation, or damages in body organs such as inblood vessels, heart, lungs, skin, and muscles.

In one embodiment, provided herein is a method of treating, preventingand/or managing psoriasis. As used herein, “psoriasis” encompasses alltypes and manifestations of psoriasis. Examples include, but are notlimited to, plaque psoriasis (e.g., chronic plaque psoriasis, moderateplaque psoriasis and severe plaque psoriasis), guttate psoriasis,inverse psoriasis, pustular psoriasis, pemphigus vulgaris, erythrodermicpsoriasis, psoriasis associated with inflammatory bowel disease (IBD),and psoriasis associated with rheumatoid arthritis (RA). Also providedherein is a method of treating, preventing and/or managing one or moresymptoms associated with psoriasis. Examples of the symptoms include,but are not limited to: red patches of skin covered with silvery scales;small scaling spots; dry, cracked skin that may bleed; itching; burning;soreness; thickened, pitted or ridged nails; and swollen and stiffjoints.

In one embodiment, provided herein is a method of treating, preventingand/or managing fibrosis and fibrotic condition. As used herein,“fibrosis” or “fibrotic condition encompasses all types andmanifestations of fibrosis or fibrotic condition. Examples include, butare not limited to, formation or deposition of tissue fibrosis; reducingthe size, cellularity (e.g., fibroblast or immune cell numbers),composition; or cellular content, of a fibrotic lesion; reducing thecollagen or hydroxyproline content, of a fibrotic lesion; reducingexpression or activity of a fibrogenic protein; reducing fibrosisassociated with an inflammatory response; decreasing weight lossassociated with fibrosis; or increasing survival.

In certain embodiments, the fibrotic condition is primary fibrosis. Inone embodiment, the fibrotic condition is idiopathic. In otherembodiments, the fibrotic condition is associated with (e.g., issecondary to) a disease (e.g., an infectious disease, an inflammatorydisease, an autoimmune disease, a malignant or cancerous disease, and/ora connective disease); a toxin; an insult (e.g., an environmental hazard(e.g., asbestos, coal dust, polycyclic aromatic hydrocarbons), cigarettesmoking, a wound); a medical treatment (e.g., surgical incision,chemotherapy or radiation), or a combination thereof.

In some embodiments, the fibrotic condition is associated with anautoimmune disease selected from scleroderma or lupus, e.g., systemiclupus erythematosus. In some embodiments, the fibrotic condition issystemic. In some embodiments, the fibrotic condition is systemicsclerosis (e.g., limited systemic sclerosis, diffuse systemic sclerosis,or systemic sclerosis sine scleroderma), nephrogenic systemic fibrosis,cystic fibrosis, chronic graft vs. host disease, or atherosclerosis.

In certain embodiments, the fibrotic condition is a fibrotic conditionof the lung, a fibrotic condition of the liver, a fibrotic condition ofthe heart or vasculature, a fibrotic condition of the kidney, a fibroticcondition of the skin, a fibrotic condition of the gastrointestinaltract, a fibrotic condition of the bone marrow or a hematopoietictissue, a fibrotic condition of the nervous system, a fibrotic conditionof the eye, or a combination thereof.

In other embodiment, the fibrotic condition affects a tissue chosen fromone or more of muscle, tendon, cartilage, skin (e.g., skin epidermis orendodermis), cardiac tissue, vascular tissue (e.g., artery, vein),pancreatic tissue, lung tissue, liver tissue, kidney tissue, uterinetissue, ovarian tissue, neural tissue, testicular tissue, peritonealtissue, colon, small intestine, biliary tract, gut, bone marrow,hematopoietic tissue, or eye (e.g., retinal) tissue.

In some embodiments, the fibrotic condition is a fibrotic condition ofthe eye. In some embodiments, the fibrotic condition is glaucoma,macular degeneration (e.g., age-related macular degeneration), macularedema (e.g., diabetic macular edema), retinopathy (e.g., diabeticretinopathy), or dry eye disease.

In certain embodiments, the fibrotic condition is a fibrotic conditionof the lung. In certain embodiments, the fibrotic condition of the lungis chosen from one or more of: pulmonary fibrosis, idiopathic pulmonaryfibrosis (IPF), usual interstitial pneumonitis (UIP), interstitial lungdisease, cryptogenic fibrosing alveolitis (CFA), bronchiectasis, andscleroderma lung disease. In one embodiment, the fibrosis of the lung issecondary to a disease, a toxin, an insult, a medical treatment, or acombination thereof. For example, the fibrosis of the lung can beassociated with (e.g., secondary to) one or more of: a disease processsuch as asbestosis and silicosis; an occupational hazard; anenvironmental pollutant; cigarette smoking; an autoimmune connectivetissue disorders (e.g., rheumatoid arthritis, scleroderma and systemiclupus erythematosus (SLE)); a connective tissue disorder such assarcoidosis; an infectious disease, e.g., infection, particularlychronic infection; a medical treatment, including but not limited to,radiation therapy, and drug therapy, e.g., chemotherapy (e.g., treatmentwith as bleomycin, methotrexate, amiodarone, busulfan, and/ornitrofurantoin). In one embodiment, the fibrotic condition of the lungtreated with the methods provided herein is associated with (e.g.,secondary to) a cancer treatment, e.g., treatment of a cancer (e.g.,squamous cell carcinoma, testicular cancer, Hodgkin's disease withbleomycin). In one embodiment, the fibrotic condition of the lung isassociated with an autoimmune connective tissue disorder (e.g.,scleroderma or lupus, e.g., SLE).

In certain embodiments, the fibrotic condition is a fibrotic conditionof the liver. In certain embodiments, the fibrotic condition of theliver is chosen from one or more of: fatty liver disease, steatosis(e.g., nonalcoholic steatohepatitis (NASH), cholestatic liver disease(e.g., primary biliary cirrhosis (PBC)), cirrhosis, alcohol inducedliver fibrosis, biliary duct injury, biliary fibrosis, orcholangiopathies. In other embodiments, hepatic or liver fibrosisincludes, but is not limited to, hepatic fibrosis associated withalcoholism, viral infection, e.g., hepatitis (e.g., hepatitis C, B orD), autoimmune hepatitis, non-alcoholic fatty liver disease (NAFLD),progressive massive fibrosis, exposure to toxins or irritants (e.g.,alcohol, pharmaceutical drugs and environmental toxins).

In certain embodiments, the fibrotic condition is a fibrotic conditionof the heart. In certain embodiments, the fibrotic condition of theheart is myocardial fibrosis (e.g., myocardial fibrosis associated withradiation myocarditis, a surgical procedure complication (e.g.,myocardial post-operative fibrosis), infectious diseases (e.g., Chagasdisease, bacterial, trichinosis or fungal myocarditis)); granulomatous,metabolic storage disorders (e.g., cardiomyopathy, hemochromatosis);developmental disorders (e.g., endocardial fibroelastosis);arteriosclerotic, or exposure to toxins or irritants (e.g., drug inducedcardiomyopathy, drug induced cardiotoxicity, alcoholic cardiomyopathy,cobalt poisoning or exposure). In certain embodiments, the myocardialfibrosis is associated with an inflammatory disorder of cardiac tissue(e.g., myocardial sarcoidosis). In some embodiments, the fibroticcondition is a fibrotic condition associated with a myocardialinfarction. In some embodiments, the fibrotic condition is a fibroticcondition associated with congestive heart failure.

In certain embodiments, the fibrotic condition is a fibrotic conditionof the kidney. In certain embodiments, the fibrotic condition of thekidney is chosen from one or more of: renal fibrosis (e.g., chronickidney fibrosis), nephropathies associated with injury/fibrosis (e.g.,chronic nephropathies associated with diabetes (e.g., diabeticnephropathy)), lupus, scleroderma of the kidney, glomerular nephritis,focal segmental glomerular sclerosis, IgA nephropathyrenal fibrosisassociated with human chronic kidney disease (CKD), chronic progressivenephropathy (CPN), tubulointerstitial fibrosis, ureteral obstruction,chronic uremia, chronic interstitial nephritis, radiation nephropathy,glomerulosclerosis, progressive glomerulonephrosis (PGN),endothelial/thrombotic microangiopathy injury, HIV-associatednephropathy, or fibrosis associated with exposure to a toxin, anirritant, or a chemotherapeutic agent. In one embodiment, the fibroticcondition of the kidney is scleroderma of the kidney. In someembodiments, the fibrotic condition of the kidney is transplantnephropathy, diabetic nephropathy, lupus nephritis, or focal segmentalglomerulosclerosis (FSGS).

In certain embodiments, the fibrotic condition is a fibrotic conditionof the skin. In certain embodiments, the fibrotic condition of the skinis chosen from one or more of: skin fibrosis (e.g., hypertrophicscarring, keloid), scleroderma, nephrogenic systemic fibrosis (e.g.,resulting after exposure to gadolinium (which is frequently used as acontrast substance for MRIs) in patients with severe kidney failure),and keloid.

In certain embodiments, the fibrotic condition is a fibrotic conditionof the gastrointestinal tract. In certain embodiments, the fibroticcondition is chosen from one or more of: fibrosis associated withscleroderma; radiation induced gut fibrosis; fibrosis associated with aforegut inflammatory disorder such as Barrett's esophagus and chronicgastritis, and/or fibrosis associated with a hindgut inflammatorydisorder, such as inflammatory bowel disease (IBD), ulcerative colitisand Crohn's disease. In some embodiments, the fibrotic condition of thegastrointestinal tract is fibrosis associated with scleroderma.

In certain embodiments, the fibrotic condition is a fibrotic conditionof the bone marrow or a hematopoietic tissue. In certain embodiments,the fibrotic condition of the bone marrow is an intrinsic feature of achronic myeloproliferative neoplasm of the bone marrow, such as primarymyelofibrosis (also referred to herein as agnogenic myeloid metaplasiaor chronic idiopathic myelofibrosis). In other embodiments, the bonemarrow fibrosis is associated with (e.g., is secondary to) a malignantcondition or a condition caused by a clonal proliferative disease. Inother embodiments, the bone marrow fibrosis is associated with ahematologic disorder (e.g., a hematologic disorder chosen from one ormore of polycythemia vera, essential thrombocythemia, myelodysplasia,hairy cell leukemia, lymphoma (e.g., Hodgkin or non-Hodgkin lymphoma),multiple myeloma or chronic myelogeneous leukemia (CML)). In yet otherembodiments, the bone marrow fibrosis is associated with (e.g.,secondary to) a non-hematologic disorder (e.g., a non-hematologicdisorder chosen from solid tumor metastasis to bone marrow, anautoimmune disorder (e.g., systemic lupus erythematosus, scleroderma,mixed connective tissue disorder, or polymyositis), an infection (e.g.,tuberculosis), or secondary hyperparathyroidism associated with vitaminD deficiency. In some embodiments, the fibrotic condition is idiopathicor drug-induced myelofibrosis. In some embodiments, the fibroticcondition of the bone marrow or hematopoietic tissue is associated withsystemic lupus erythematosus or scleroderma.

In one embodiment, provided herein is a method of treating, preventingand/or managing scleroderma. Scleroderma is a group of diseases thatinvolve hardening and tightening of the skin and/or other connectivetissues. Scleroderma may be localized (e.g., affecting only the skin) orsystemic (e.g., affecting other systems such as, e.g., blood vesselsand/or internal organs). Common symptoms of scleroderma includeRaynaud's phenomenon, gastroesophageal reflux disease, and skin changes(e.g., swollen fingers and hands, or thickened patches of skin). In someembodiments, the scleroderma is localized, e.g., morphea or linearscleroderma. In some embodiments, the condition is a systemic sclerosis,e.g., limited systemic sclerosis, diffuse systemic sclerosis, orsystemic sclerosis sine scleroderma.

Localized scleroderma (localized cutaneous fibrosis) includes morpheaand linear scleroderma. Morphea is typically characterized byoval-shaped thickened patches of skin that are white in the middle, witha purple border. Linear scleroderma is more common in children. Symptomsof linear scleroderma may appear mostly on one side of the body. Inlinear scleroderma, bands or streaks of hardened skin may develop on oneor both arms or legs or on the forehead. En coup de sabre (frontallinear scleroderma or morphea en coup de sabre) is a type of localizedscleroderma typically characterized by linear lesions of the scalp orface.

Systemic scleroderma (systemic sclerosis) includes, e.g., limitedsystemic sclerosis (also known as limited cutaneous systemic sclerosis,or CREST syndrome), diffuse systemic sclerosis (also known as diffusecutaneous systemic sclerosis), and systemic sclerosis sine scleroderma.CREST stands for the following complications that may accompany limitedscleroderma: calcinosis (e.g., of the digits), Raynaud's phenomenon,esophageal dysfunction, sclerodactyly, and telangiectasias. Typically,limited scleroderma involves cutaneous manifestations that mainly affectthe hands, arms, and face. Limited and diffuse subtypes aredistinguished based on the extent of skin involvement, with sparing ofthe proximal limbs and trunk in limited disease. See, e.g., Denton, C.P. et al. (2006), Nature Clinical Practice Rheumatology, 2(3):134-143.The limited subtype also typically involves a long previous history ofRaynaud's phenomenon, whereas in the diffuse subtype, onset of Raynaud'sphenomenon can be simultaneous with other manifestations or might occurlater. Both limited and diffuse subtypes may involve internal organs.Typical visceral manifestations of limited systemic sclerosis includeisolated pulmonary hypertension, severe bowel involvement, and pulmonaryfibrosis. Typical visceral manifestations of diffuse systemic sclerosisinclude renal crisis, lung fibrosis, and cardiac disease. Diffusesystemic sclerosis typically progresses rapidly and affects a large areaof the skin and one or more internal organs (e.g., kidneys, esophagus,heart, or lungs). Systemic sclerosis sine scleroderma is a rare disorderin which patients develop vascular and fibrotic damage to internalorgans in the absence of cutaneous sclerosis.

In one embodiment, provided herein is a method of treating, preventingand/or managing inflammatory myopathies. As used herein, “inflammatorymyopathies” encompass all types and manifestations of inflammatorymyopathies. Examples include, but are not limited to, muscle weakness(e.g., proximal muscle weakness), skin rash, fatigue after walking orstanding, tripping or falling, dysphagia, dysphonia, difficultybreathing, muscle pain, tender muscles, weight loss, low-grade fever,inflamed lungs, light sensitivity, calcium deposits (calcinosis) underthe skin or in the muscle, as well as biological concomitants ofinflammatory myopathies as disclosed herein or as known in the art.Biological concomitants of inflammatory myopathies (e.g.,dermatomyositis) include, e.g., altered (e.g., increased) levels ofcytokines (e.g., Type I interferons (e.g., IFN-α and/or IFN-β),interleukins (e.g., IL-6, IL-10, IL-15, IL-17 and IL-18), and TNF-α),TGF-β, B-cell activating factor (BAFF), overexpression of IFN induciblegenes (e.g., Type I IFN inducible genes). Other biological concomitantsof inflammatory myopathies can include, e.g., an increased erythrocytesedimentation rate (ESR) and/or elevated level of creatine kinase.Further biological concomitants of inflammatory myopathies can includeautoantibodies, e.g., anti-synthetase autoantibodies (e.g., anti-Jo1antibodies), anti-signal recognition particle antibodies (anti-SRP),anti-Mi-2 antibodies, anti-p155 antibodies, anti-PM/Sci antibodies, andanti-RNP antibodies.

The inflammatory myopathy can be an acute inflammatory myopathy or achronic inflammatory myopathy. In some embodiments, the inflammatorymyopathy is a chronic inflammatory myopathy (e.g., dermatomyositis,polymyositis, or inclusion body myositis). In some embodiments, theinflammatory myopathy is caused by an allergic reaction, another disease(e.g., cancer or a connective tissue disease), exposure to a toxicsubstance, a medicine, or an infectious agent (e.g., a virus). In someembodiments, the inflammatory myopathy is associated with lupus,rheumatoid arthritis, or systemic sclerosis. In some embodiments, theinflammatory myopathy is idiopathic. In some embodiments, theinflammatory myopathy is selected from polymyositis, dermatomyositis,inclusion body myositis, and immune-mediated necrotizing myopathy. Insome embodiments, the inflammatory myopathy is dermatomyositis.

In another embodiment, provided herein is a method of treating,preventing and/or managing a skin condition (e.g., a dermatitis). Insome embodiments, the methods provided herein can reduce symptomsassociated with a skin condition (e.g., itchiness and/or inflammation).In some such embodiments, the compound provided herein is administeredtopically (e.g., as a topical cream, eye-drop, nose drop or nasalspray). In some such embodiments, the compound is a PI3K delta inhibitor(e.g., a PI3K inhibitor that demonstrates greater inhibition of PI3Kdelta than of other PI3K isoforms). In some embodiments, the PI3K deltainhibitor prevents mast cell degranulation.

As used herein, “skin condition” includes any inflammatory condition ofthe skin (e.g., eczema or dermatitis, e.g., contact dermatitis, atopicdermatitis, dermatitis herpetiformis, seborrheic dermatitis, nummulardermatitis, stasis dermatitis, perioral dermatitis), as well asaccompanying symptoms (e.g., skin rash, itchiness (pruritis), swelling(edema), hay fever, anaphalaxis). Frequently, such skin conditions arecaused by an allergen. As used herein, a “skin condition” also includes,e.g., skin rashes (e.g., allergic rashes, e.g., rashes resulting fromexposure to allergens such as poison ivy, poison oak, or poison sumac,or rashes caused by other diseases or conditions), insect bites, minorburns, sunburn, minor cuts, and scrapes. In some embodiments, thesymptom associated with inflammatory myopathy, or the skin condition orsymptom associated with the skin condition, is a skin rash or itchiness(pruritis) caused by a skin rash.

The skin condition (e.g., the skin rash) may be spontaneous, or it maybe induced, e.g., by exposure to an allergen (e.g., poison ivy, poisonoak, or poison sumac), drugs, food, insect bite, inhalants, emotionalstress, exposure to heat, exposure to cold, or exercise. In someembodiments, the skin condition is a skin rash (e.g., a pruritic rash,e.g., utricaria). In some embodiments, the skin condition is an insectbite. In some embodiments, the skin condition is associated with anotherdisease (e.g., an inflammatory myopathy, e.g., dermatomyositis).

In some embodiments, the subject (e.g., the subject in need of treatmentfor an inflammatory myopathy and/or a skin condition) exhibits anelevated level or elevated activity of IFN-α, TNF-α, IL-6, IL-8, IL-1,or a combination thereof. In certain embodiments, the subject exhibitsan elevated level of IFN-α. In some embodiments, treating (e.g.,decreasing or inhibiting) the inflammatory myopathy, or the skincondition, comprises inhibiting (e.g., decreasing a level of, ordecreasing a biological activity of) one or more of IFN-α, TNF-α, IL-6,IL-8, or IL-1 in the subject or in a sample derived from the subject. Insome embodiments, the method decreases a level of IFN-α, TNF-α, IL-6,IL-8, or IL-1 in the subject or in a sample derived from the subject. Insome embodiments, the method decreases a level of IFN-α in the subjector in a sample derived from the subject. In some embodiments, the levelof IFN-α, TNF-α, IL-6, IL-8, or IL-1 is the level assessed in a sampleof whole blood or PBMCs. In some embodiments, the level of IFN-α, TNF-α,IL-6, IL-8, or IL-1 is the level assessed in a sample obtained by a skinbiopsy or a muscle biopsy. In some embodiments, the sample is obtainedby a skin biopsy.

In one embodiment, provided herein is a method of treating, preventingand/or managing myositis. As used herein, “myositis” encompasses alltypes and manifestations of myositis. Examples include, but are notlimited to, myositis ossificans, fibromyositis, idiopathic inflammatorymyopathies, dermatomyositis, juvenile dermatomyositis, polymyositis,inclusion body myositis and pyomyositis. In one embodiment, the diseaseor disorder is dermatomyositis. Also provided herein is a method oftreating, preventing and/or managing one or more symptoms associatedwith myositis. Examples of the symptoms include, but are not limited to:muscle weakness; trouble lifting arms; trouble swallowing or breathing;muscle pain; muscle tenderness; fatigue; fever; lung problems;gastrointestinal ulcers; intestinal perforations; calcinosis under theskin; soreness; arthritis; weight loss; and rashes.

In one embodiment, provided herein is a method of treating, preventingand/or managing lupus. As used herein, “lupus” refers to all types andmanifestations of lupus. Examples include, but are not limited to,systemic lupus erythematosus; lupus nephritis; cutaneous manifestations(e.g., manifestations seen in cutaneous lupus erythematosus, e.g., askin lesion or rash); CNS lupus; cardiovascular, pulmonary, hepatic,hematological, gastrointestinal and musculoskeletal manifestations;neonatal lupus erythematosus; childhood systemic lupus erythematosus;drug-induced lupus erythematosus; anti-phospholipid syndrome; andcomplement deficiency syndromes resulting in lupus manifestations. Inone embodiment, the lupus is systemic lupus erythematosus (SLE),cutaneous lupus erythematosus (CLE), drug-induced lupus, or neonatallupus. In another embodiment, the lupus is a CLE, e.g., acute cutaneouslupus erythematosus (ACLE), subacute cutaneous lupus erythematosus(SCLE), intermittent cutaneous lupus erythematosus (also known as lupuserythematosus tumidus (LET)), or chronic cutaneous lupus. In someembodiments, the intermittent CLE is chronic discloid lupuserythematosus (CDLE) or lupus erythematosus profundus (LEP) (also knownas lupus erythematosus panniculitis). Types, symptoms, and pathogenesisof CLE are described, for example, in Wenzel et al. (2010), Lupus, 19,1020-1028.

In one embodiment, provided herein is a method of treating, preventingand/or managing Sjögren's syndrome. As used herein, “Sjögren's syndrome”refers to all types and manifestations of Sjögren's syndrome. Examplesinclude, but are not limited to, primary and secondary Sjögren'ssyndrome. Also provided herein is a method of treating, preventingand/or managing one or more symptoms associated with Sjögren's syndrome.Examples of the symptoms include, but are not limited to: dry eyes; drymouth; joint pain; swelling; stiffness; swollen salivary glands; skinrashes; dry skin; vaginal dryness; persistent dry cough; and prolongedfatigue.

In some embodiments, provided herein is a method of treating a bonedisorder in a subject that comprises administering to said subject atherapeutically effective amount of a compound provided herein (e.g., aPI3K-γ selective compound provided herein), or a pharmaceuticallyacceptable form (e.g., pharmaceutically acceptable salts, hydrates,solvates, isomers, prodrugs, and isotopically labeled derivatives)thereof, or a pharmaceutical composition as provided herein. As usedherein, “bone disorder” encompasses all types and manifestations of bonedisorders. Exemplary bone disorders include, but are not limited to,bone cancer, bone metastases, osteoporosis, fracture repair, avascularnecrosis (osteonecrosis), bone spur (osteophytes), craniosynostosis,Coffin-Lowry syndrome, fibrodysplasia ossificans progressive, fibrousdysplasia, Fong Disease (Nail-patella syndrome), Giant cell tumor ofbone, Greenstick Fracture, hypophosphatasia, Klippel-Feil syndrome,metabolic bone disease, osteoarthritis, osteitis deformans (Paget'sdisease of bone), osteitis fibrosa cystica (osteitis fibrosa or VonRecklinghausen's disease of bone), osteitis pubis, condensing osteitis(osteitis condensas), osteochondritis dissecans, osteochondroma (bonetumor), osteogenesis imperfect, osteomalacia, osteomyelitis, osteopenia,osteopetrosis, porotic hyperostosis, primary hyperparathyroidism, renalosteodystrophy, Salter-Harris fractures, and water on the knee. In oneembodiment, the bone disorder is a systemical bone disorder. In anotherembodiment, the bone disorder is a topical bone disorder. In oneembodiment, the bone disorder is associated with excess bone formation.In another embodiment, the bone disorder is associated with excess boneresorption. In one embodiment, without being limited by a particulartheory, a compound provided herein inhibits differentiation ofosteoclasts from bone marrow macrophages.

In some embodiments, a symptom associated with the disease or disorderprovided herein is reduced by at least 10%, at least 20%, at least 30%,at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, atleast 90%, or at least 95% relative to a control level. The controllevel includes any appropriate control as known in the art. For example,the control level can be the pre-treatment level in the sample orsubject treated, or it can be the level in a control population (e.g.,the level in subjects who do not have the disease or disorder or thelevel in samples derived from subjects who do not have the disease ordisorder). In some embodiments, the decrease is statisticallysignificant, for example, as assessed using an appropriate parametric ornon-parametric statistical comparison.

Methods of Treatment, Prevention and/or Management for Pulmonary orRespiratory Disorders

Without being limited by a particular theory, it was found thatadministering a compound provided herein (e.g., Compound 4) byinhalation can accord various therapeutic benefits as described hereinin treating, preventing and/or managing pulmonary or respiratorydiseases. Accordingly, in certain embodiments, provided herein is amethod of treating, preventing, and/or managing a pulmonary orrespiratory disease in a subject, comprising administering to a subjectin need thereof by inhalation a therapeutically or prophylacticallyeffective amount of a compound provided herein, or an enantiomer, amixture of enantiomers, or a mixture of two or more diastereomersthereof, or a pharmaceutically acceptable form thereof.

In addition, without being limited by a particular theory, it was foundthat administering a compound provided herein by inhalation results in aprolonged retainment of the compound in patient's lung. Thus, in someembodiments, provided herein is a method of eliciting prolongedanti-inflammatory effect in lung in a subject suffering from a pulmonaryor respiratory disease, comprising administering to the subject byinhalation a therapeutically or prophylactically effective amount of acompound provided herein, or an enantiomer, a mixture of enantiomers, ora mixture of two or more diastereomers thereof, or a pharmaceuticallyacceptable form thereof, wherein the compound is retained in lung for aprolonged period (e.g., a period longer than what is provided by oraladministration).

In some embodiments, the compound is retained in lung for about hour,about 3 hours, about 6 hours, about 12 hours, about 24 hours, about 48hours, or about 72 hours longer than what is provided by oraladministration.

In some embodiments, more than 80%, more than 70%, more than 60%, morethan 50%, more than 40%, more than 30%, or more than 20% of the amountof the compound as initially administered to patient remains in lung at24 hours after administration by inhalation.

In some embodiments, the concentration of the compound in lung followingadministration by inhalation is about 100, about 200, about 500, about1000, about 2000, about 3000, about 4000, about 5000, about 6000, about7000, about 8000, about 9000, or about 10000 times higher than theplasma concentration of the compound at about 5 hours after theadministration. In some embodiments, the concentration of the compoundin lung following administration by inhalation is about 100, about 200,about 500, about 1000, about 2000, about 3000, about 4000, about 5000,about 6000, about 7000, about 8000, about 9000, or about 10000 timeshigher than the plasma concentration of the compound at about 12 hoursafter the administration. In some embodiments, the concentration of thecompound in lung following administration by inhalation is about 100,about 200, about 500, about 1000, about 2000, about 3000, about 4000,about 5000, about 6000, about 7000, about 8000, about 9000, or about10000 times higher than the plasma concentration of the compound atabout 24 hours after the administration.

In some embodiments, the compound is administered at a dose of less than0.01 μg/kg/day, less than 0.02 μg/kg/day, less than 0.05 μg/kg/day, lessthan 0.1 μg/kg/day, less than 0.2 μg/kg/day, less than 0.5 μg/kg/day,less than 1 μg/kg/day, less than 2 μg/kg/day, less than 5 μg/kg/day,less than 10 μg/kg/day, less than 20 μg/kg/day, less than 50 μg/kg/day,or less than 100 μg/kg/day. In some embodiments, the compound isadministered at a dose of about 0.01 μg/kg/day, about 0.02 μg/kg/day,about 0.05 μg/kg/day, about 0.1 μg/kg/day, about 0.2 μg/kg/day, about0.5 μg/kg/day, about 1 μg/kg/day, about 2 μg/kg/day, about 5 μg/kg/day,about 10 μg/kg/day, about 20 μg/kg/day, about 50 μg/kg/day, or about 100μg/kg/day. In some embodiments, the compound is administered at a doseof from about 0.01 μg/kg/day to about 100 μg/kg/day, from about 0.01μg/kg/day to about 50 μg/kg/day, from about 0.01 μg/kg/day to about 20μg/kg/day, from about 0.01 μg/kg/day to about 10 μg/kg/day, from about0.01 μg/kg/day to about 5 μg/kg/day, from about 0.01 μg/kg/day to about1 μg/kg/day, from about 0.05 μg/kg/day to about 1 μg/kg/day, or fromabout 0.1 μg/kg/day to about 1 μg/kg/day.

In one embodiment, the compound is administered once daily (QD). Inanother embodiment, the compound is administered twice daily (BID). Inanother embodiment, the compound is administered three time daily (TID).In another embodiment, the compound is administered four times daily(QID).

In one embodiment, the subject is a mammal. In another embodiment, thesubject is a human.

In one embodiment, provided herein is a method of treating, preventing,and/or managing a pulmonary or respiratory disease in a subject,comprising administering to a subject in need thereof by inhalation atherapeutically or prophylactically effective amount of a PI3Kγinhibitor, or an enantiomer, a mixture of enantiomers, or a mixture oftwo or more diastereomers thereof, or a pharmaceutically acceptable formthereof. In some embodiments, the PI3Kγ inhibitor has a delta/gammaselectivity ratio of greater than about 1 to <10, greater than about 10to <50, or greater than about 50 to <350. In some embodiments, the PI3Kγinhibitor has a delta/gamma selectivity ratio of greater than about 1,greater than about 5, greater than about 10, greater than about 15,greater than about 20, greater than about 25, greater than about 50,greater than about 75, greater than about 100, greater than about 150,greater than about 200, greater than about 250, greater than about 300,greater than about 350, greater than about 500, or greater than about1000.

In one embodiment, provided herein is a method of treating, preventing,and/or managing a pulmonary or respiratory disease in a subject,comprising administering to a subject in need thereof by inhalation atherapeutically or prophylactically effective amount of Compound 4, oran enantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof. Inone embodiment, Compound 4 is a PI3Kγ inhibitor. In one embodiment, thecompound, e.g., Compound 4 has a delta/gamma selectivity ratio ofgreater than about 1 to <10, greater than about 10 to <50, or greaterthan about 50 to <350. In one embodiment, the compound, e.g., Compound 4has a delta/gamma selectivity ratio of greater than about 50 to <350. Inone embodiment, the compound, e.g., Compound 4 has a delta/gammaselectivity ratio of greater than about 50 to <150. In one embodiment,the compound, e.g., Compound 4 has a delta/gamma selectivity ratio ofgreater than about 75 to <125. In one embodiment, the compound has adelta/gamma selectivity ratio of greater than about 100. In someembodiments, the compound has a delta/gamma selectivity ratio of greaterthan about 1, greater than about 5, greater than about 10, greater thanabout 15, greater than about 20, greater than about 25, greater thanabout 50, greater than about 75, greater than about 100, greater thanabout 150, greater than about 200, greater than about 250, greater thanabout 300, greater than about 350, greater than about 500, or greaterthan about 1000.

Administration by Inhalation

Many diseases of the respiratory tract are known to respond to treatmentby the direct application of therapeutic agents by inhalation. Suchadministration can result in the better utilization of the medicament inthat the drug is deposited directly at the desired site and where itsaction may be required. Therefore, without being limited by a particulartheory, administration by inhalation can significantly reduce the doserequired to achieve therapeutic efficacy, which, in turn can result inmarked reduction of undesired side effects and cost of medicament. It istypically accepted in the industry that the bioavailability of the drugis optimum when the drug particles delivered to the respiratory tractare between 1 to 5 microns in size.

Various methods and devices can be used to deliver a compound providedherein by inhalation. The inhalable formulation can be administered viathe mouth or nose ultimately for pulmonary delivery thereof. Forexample, dry powder inhalers (DPIs), which usually have a means forintroducing the drug (active drug plus carrier) into a high velocity airstream, can be used to practice the methods provided herein. The highvelocity air stream is used as the primary mechanism for breaking up thecluster of micronized particles or separating the drug particles fromthe carrier. Inhalation devices useful for dispensing powder forms ofmedicament such as those described in U.S. Pat. Nos. 3,507,277;3,518,992; 3,635,219; 3,795,244; and 3,807,400, are encompassed by thecurrent disclosure. In certain embodiments, such devices also includepropeller means, which upon inhalation aid in dispensing the powder outof the capsule, so that it is not necessary to rely solely on theinhaled air to suction powder from the capsule. (See, e.g., U.S. Pat.Nos. 2,517,482; 3,831,606; 3,948,264; and 5,458, 135, all of which areincorporated herein by reference). In certain embodiments, utilizationof vibration to facilitate suspension of power into an inhaled gasstream and which utilizes synthetic jetting to aerosolize drug powderfrom a blister pack is also provided herein. (See, e.g., U.S. Pat. Nos.7,318,434 and 7,334,577, incorporated herein by reference). In someembodiments, controlled aliquots or doses of a medication orpre-packaged drug in a blister pack, which includes a frangible crownedtop element which can be conical, conical with a rounded point, rounded,such as those described in U.S. Pat. No. 7,080,644, are alsoencompassed.

In certain embodiments, a compound provided herein is administered usingmetered dose inhalers (MDIs). MDIs typically have a pressurized canisterfilled with a liquid propellant. The drug is either suspended ordissolved in the propellant. The MDIs have a metering valve for meteringout a known quantity of the propellant and hence the drug. When thecanister is depressed against the MDI housing a known quantity of thepropellant is discharged. The propellant evaporates leaving behind afine aerosol of the drug suitable for inhalation by the patient. Incertain embodiments, MDIs that contain a breath actuation mechanism aspacer are also encompassed herein.

In some embodiments, a compound provided herein is administered usingnebulizers, such as the jet nebulizers. Nebulizers produce a fineaerosol mist/droplets which carry the drug either as a suspension ordissolved in the aqueous medium. The jet nebulizers use compressed airto atomize the aqueous solution. A drug can be administered to a patientwith repetitive non-forced inhalation over a prolonged period of time.

Examples of devices suitable for such pulmonary delivery include, butare not limited to, air-jet, ultrasonic, or vibrating-mesh devices suchas Pari LC Star, Aeroeclipse II, Prodose (HaloLite), Acorn II, TUp-draft II, Sidestream, AeroTech II, Mini heart, MisterNeb, Sonix 2000,MABISMist II and other suitable aerosol systems. In some embodiments,the nebulizer is a vibrating-mesh nebulizer that could include anAERONEB PRO, AERONEB SOLO, AERONEB GO, AERONEB LAB, OMRON MICROAIR, PARTEFLOW, RESPIRONICS I-NEB, or other suitable devices.

Pulmonary or Respiratory Diseases

Provided herein is a method of treating, preventing, and/or managingpulmonary or respiratory disease using a compound provided herein.Examples of pulmonary or respiratory disease include, but are notlimited to, lung inflammation, chronic obstructive pulmonary disease,asthma, pneumonia, hypersensitivity pneumonitis, pulmonary infiltratewith eosinophilia, environmental lung disease, pneumonia,bronchiectasis, cystic fibrosis, interstitial lung disease, postinflammatory pulmonary fibrosis, primary pulmonary hypertension,pulmonary thromboembolism, disorders of the pleura, disorders of themediastinum, disorders of the diaphragm, disorders of the larynx,disorders of the trachea, acute lung injury, hypoventilation,hyperventilation, sleep apnea, acute respiratory distress syndrome,mesothelioma, sarcoma, graft rejection, graft versus host disease, lungcancer, allergic rhinitis, allergy, allergic bronchopulmonaryaspergillosis, asbestosis, aspergilloma, aspergillosis, bronchiectasis,chronic bronchitis, emphysema, eosinophilic pneumonia, idiopathicpulmonary fibrosis, idiopathic interstitial pneumonia, non-specificinterstitial pneumonia (NSIP), bronchiolitis obliterans with organizingpneumonia (BOOP, also called cryptogenic organizing pneumonia or COP),lymphocytic interstitial pneumonia (LIP), acute interstitial pneumonitisinvasive pneumococcal disease, pneumococcal pneumonia, influenza,nontuberculous mycobacteria, pleural effusion, a pleural cavity disease,empyema, pleurisy, pneumoconiosis, pneumocytosis, respiratory viralinfection, acute bronchitis, aspiration pneumonia, ventilator-associatedpneumonia, pneumocystic jiroveci pneumonia, pneumonia, pulmonaryactinomycosis, pulmonary alveolar proteinosis, pulmonary anthrax,pulmonary edema, pulmonary embolus, pulmonary embolism, acute chestsyndrome, idiopathic pulmonary hemosiderosis, pulmonary hemorrhage,pulmonary hyperplasia, pulmonary inflammation, pulmonary histiocytosisX, eosinophilic granuloma, pulmonary Langerhan's cell histiocytosis,occupational lung disease, pneumopathy due to inhalation of dust,respiratory conditions due to chemical fumes and vapors, lipoidpneumonia, pulmonary hypertension, pulmonary arterial hypertension,pulmonary nocardiosis, pulmonary tuberculosis, pulmonary veno-occlusivedisease, pulmonary vascular disease, rheumatoid lung disease, connectivetissue disease-associated interstitial lung disease (e.g., systemicsclerosis (SSc or scleroderma)-associated interstitial lung disease,polymyositis-associated interstitial lung disease,dermatomyositis-associated interstitial lung disease, rheumatoidarthritis-associated interstitial lung disease, systemic lupuserythematosus-associated interstitial lung disease, interstitial lungdisease associated with Sjögren's syndrome, mixed connective tissuedisease-associated interstitial lung disease, and ankylosingspondylitis-associated interstitial lung disease), a restrictive lungdisease, a respiratory tract infection (upper and lower), sarcoidosis,Wegener's granulomatosis (also known as granulomatosis with polyangiitis(GPA) or necrotizing granulomatous vasculitis (NGV)), Churg-StraussSyndrome, microscopic polyangiitis (MPA), small cell lung carcinoma,non-small cell lung carcinoma, lymphangioleiomyomatosis (LAM),radiation-induced lung disease (also known as radiation pneumonitis),pulmonary vasculitis, viral pneumonia, pneumococcal pneumonia, bacterialpneumonia, bronchopneumonia, epithelial tumors, papillomas, adenomas,squamous cell carcinoma, small cell carcinoma, adenocarcinoma, largecell carcinoma, adenosquamous carcinoma, carcinoid tumor, carcinoma ofsalivary-gland type, soft tissue tumors, localized fibrous tumor,epithelioid hemangioendothelioma, pleuropulmonary blastoma, chondroma,calcifying fibrous pseudotumor of the pleura, congenital peribronchialmyofibroblastic tumor, diffuse pulmonary lymphangiomatosis, desmoplasticsmall round cell tumor, mesothelial tumors, adenomatoid tumor,epithelioid mesothelioma, sarcomatoid mesothelioma, biphasicmesothelioma, hamartoma, sclerosing hemangioma, clear cell tumor, germcell neoplasms, thymona, melanoma, and secondary tumor. In certainembodiments, provided herein is a method of treating, preventing, and/ormanaging a lymphoproliferative disease using a compound provided herein.Examples of lymphoproliferative disease include, but are not limited to,lymphoid interstitial pneumonia, nodular lymphoid hyperplasia, andlymphomatoid granulomatosis.

In certain embodiments, the pulmonary or respiratory disease to betreated, prevented and/or managed using a compound provided herein is anobstructive lung disease or disorder. In some embodiments, theobstructive lung disease is acute respiratory distress syndrome (ARDS),asthma, bronchiectasis, bronchiolectasis, bronchiolitis, bronchitis,chronic obstructive pulmonary disease (COPD), or emphysema.

In certain embodiments, in treating, preventing and/or managing apulmonary or respiratory disease provided herein, a therapeutically orprophylactically effective amount of a compound provided herein (e.g.,Compound 4), or an enantiomer, a mixture of enantiomers, or a mixture oftwo or more diastereomers thereof, or a pharmaceutically acceptable formthereof, is from about 0.005 to about 1,000 mg per day, from about 0.01to about 500 mg per day, from about 0.01 to about 250 mg per day, fromabout 0.01 to about 100 mg per day, from about 0.1 to about 100 mg perday, from about 0.5 to about 100 mg per day, from about 1 to about 100mg per day, from about 0.01 to about 50 mg per day, from about 0.1 toabout 50 mg per day, from about 0.5 to about 50 mg per day, from about 1to about 50 mg per day, from about 2 to about 25 mg per day, or fromabout 5 to about 10 mg per day.

In certain embodiments, the therapeutically or prophylacticallyeffective amount is about 0.1, about 0.2, about 0.5, about 1, about 2,about 5, about 10, about 15, about 20, about 25, about 30, about 40,about 45, about 50, about 60, about 70, about 80, about 90, about 100,or about 150 mg per day.

In one embodiment, the recommended daily dose range of a compoundprovided herein (e.g., Compound 4), or an enantiomer, a mixture ofenantiomers, or a mixture of two or more diastereomers thereof, or apharmaceutically acceptable form thereof, for the conditions describedherein lie within the range of from about 0.5 mg to about 50 mg per day,in a single once-a-day dose or in divided doses throughout a day. Insome embodiments, the dosage ranges from about 1 mg to about 50 mg perday. In some embodiments, the dosage ranges from about 0.5 to about 5 mgper day. Specific doses per day include 0.1, 0.2, 0.5, 1, 2, 3, 4, 5, 6,7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25,26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43,44, 45, 46, 47, 48, 49 or 50 mg per day.

In one embodiment, a compound provided herein (e.g., Compound 4), or anenantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof, isadministered at a dose of less than 0.1, about 0.1, less than 0.5, about0.5, between about 0.1 and about 1.0, between about 0.5 and about 1.0,about 1, or about 2 mg per day.

In another embodiment, a compound provided herein (e.g., Compound 4), oran enantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof, isadministered at a dose of less than 0.2, about 0.2, less than 1.0, about1.0, between about 0.2 and about 2.0, between about 1.0 and about 2.0,about 2, or about 4 mg per day.

In one embodiment, the dose is less than 0.1 mg per day.

In another embodiment, the dose is about 0.1 mg per day.

In another embodiment, the dose is less than 0.5 mg per day.

In another embodiment, the dose is about 0.5 mg per day.

In another embodiment, the dose is between about 0.1 and about 1.0 mgper day.

In another embodiment, the dose is between about 0.5 and about 1.0 mgper day.

In another embodiment, the dose is about 1 mg per day.

In another embodiment, the dose is about 2 mg per day.

In another embodiment, the dose is less than 0.2 mg per day.

In another embodiment, the dose is about 0.2 mg per day.

In another embodiment, the dose is less than 1.0 mg per day.

In another embodiment, the dose is about 1.0 mg per day.

In another embodiment, the dose is between about 0.2 and about 2.0 mgper day.

In another embodiment, the dose is between about 1.0 and about 2.0 mgper day.

In another embodiment, the dose is about 2 mg per day.

In another embodiment, the dose is about 4 mg per day.

In a specific embodiment, the recommended starting dosage can be 0.5, 1,2, 3, 4, 5, 10, 15, 20, 25 or 50 mg per day. In another embodiment, therecommended starting dosage can be 0.5, 1, 2, 3, 4, or 5 mg per day. Thedose can be escalated to 15, 20, 25, 30, 35, 40, 45 and 50 mg/day.

In certain embodiments, the therapeutically or prophylacticallyeffective amount is from about 0.001 to about 100 mg/kg/day, from about0.01 to about 50 mg/kg/day, from about 0.01 to about 25 mg/kg/day, fromabout 0.01 to about 10 mg/kg/day, from about 0.01 to about 9 mg/kg/day,0.01 to about 8 mg/kg/day, from about 0.01 to about 7 mg/kg/day, fromabout 0.01 to about 6 mg/kg/day, from about 0.01 to about 5 mg/kg/day,from about 0.01 to about 4 mg/kg/day, from about 0.01 to about 3mg/kg/day, from about 0.01 to about 2 mg/kg/day, or from about 0.01 toabout 1 mg/kg/day.

In certain embodiments, the therapeutically or prophylacticallyeffective amount is from about 0.01 to about 25 mg/kg/day, from about0.01 to about 20 mg/kg/day, from about 0.01 to about 15 mg/kg/day, fromabout 0.01 to about 10 mg/kg/day, from about 0.05 to about 25 mg/kg/day,from about 0.05 to about 20 mg/kg/day, from about 0.05 to about 15mg/kg/day, or from about 0.05 to about 10 mg/kg/day.

The administered dose can also be expressed in units other thanmg/kg/day. For example, doses for parenteral administration can beexpressed as mg/m²/day. One of ordinary skill in the art would readilyknow how to convert doses from mg/kg/day to mg/m²/day to given eitherthe height or weight of a subject or both (see,www.fda.gov/cder/cancer/animalframe.htm). For example, a dose of 1mg/kg/day for a 65 kg human is approximately equal to 38 mg/m²/day.

A compound provided herein (e.g., Compound 4), or an enantiomer, amixture of enantiomers, or a mixture of two or more diastereomersthereof, or a pharmaceutically acceptable form thereof, can beadministered once daily (QD), or divided into multiple daily doses suchas twice daily (BID), three times daily (TID), and four times daily(QID). In addition, the administration can be continuous (i.e., dailyfor consecutive days or every day), intermittent, e.g., in cycles (i.e.,including days, weeks, or months of rest without drug). As used herein,the term “daily” is intended to mean that a therapeutic compound, suchas Compound 4, is administered once or more than once each day, forexample, for a period of time. The term “continuous” is intended to meanthat a therapeutic compound, such as Compound 4, is administered dailyfor an uninterrupted period of at least 10 days to 52 weeks. The term“intermittent” or “intermittently” as used herein is intended to meanstopping and starting at either regular or irregular intervals. Forexample, intermittent administration of Compound 4 is administration forone to six days per week, administration in cycles (e.g., dailyadministration for two to eight consecutive weeks, then a rest periodwith no administration for up to one week), or administration onalternate days. The term “cycling” as used herein is intended to meanthat a therapeutic compound, such as Compound 4, is administered dailyor continuously but with a rest period.

In some embodiments, the frequency of administration is in the range ofabout a daily dose to about a monthly dose. In certain embodiments,administration is once a day, twice a day, three times a day, four timesa day, once every other day, twice a week, once every week, once everytwo weeks, once every three weeks, or once every four weeks. In oneembodiment, the compound provided herein is administered once a day. Inanother embodiment, the compound provided herein is administered twice aday. In yet another embodiment, the compound provided herein isadministered three times a day. In still another embodiment, thecompound provided herein is administered four times a day.

In one embodiment, a compound provided herein (e.g., Compound 4), or anenantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof, isadministered twice per day (BID). In one embodiment, the dose is about0.1, 0.2, 0.25, 0.5, 1, 2, 2.5, 5, 10, 15, 20, 25, or 50 mg BID.

In one embodiment, a compound provided herein (e.g., Compound 4), or anenantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof, isadministered at a dose of less than 0.1, about 0.1, less than 0.5, about0.5, between about 0.1 and about 1.0, between about 0.5 and about 1.0,about 1, or about 2 mg BID.

In another embodiment, a compound provided herein (e.g., Compound 4), oran enantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof, isadministered at a dose of less than 0.2, about 0.2, less than 1.0, about1.0, between about 0.2 and about 2.0, between about 1.0 and about 2.0,about 2, or about 4 mg BID.

In one embodiment, the dose is less than 0.1 mg BID.

In another embodiment, the dose is about 0.1 mg BID.

In another embodiment, the dose is less than 0.5 mg BID.

In another embodiment, the dose is about 0.5 mg BID.

In another embodiment, the dose is between about 0.1 and about 1.0 mgBID.

In another embodiment, the dose is between about 0.5 and about 1.0 mgBID.

In another embodiment, the dose is about 1 mg BID.

In another embodiment, the dose is about 2 mg BID.

In another embodiment, the dose is less than 0.2 mg BID.

In another embodiment, the dose is about 0.2 mg BID.

In another embodiment, the dose is less than 1.0 mg BID.

In another embodiment, the dose is about 1.0 mg BID.

In another embodiment, the dose is between about 0.2 and about 2.0 mgBID.

In another embodiment, the dose is between about 1.0 and about 2.0 mgBID.

In another embodiment, the dose is about 2 mg BID.

In another embodiment, the dose is about 4 mg BID.

In one embodiment, a compound provided herein (e.g., Compound 4), or anenantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof, isadministered once daily (QD). In one embodiment, the dose is about 0.1,0.2, 0.25, 0.5, 1, 2, 2.5, 5, 10, 15, 20, 25, or 50 mg QD.

In one embodiment, a compound provided herein (e.g., Compound 4), or anenantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof, isadministered at a dose of less than 0.1, about 0.1, less than 0.5, about0.5, between about 0.1 and about 1.0, between about 0.5 and about 1.0,about 1, or about 2 mg QD.

In another embodiment, a compound provided herein (e.g., Compound 4), oran enantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof, isadministered at a dose of less than 0.2, about 0.2, less than 1.0, about1.0, between about 0.2 and about 2.0, between about 1.0 and about 2.0,about 2, or about 4 mg QD.

In one embodiment, the dose is less than 0.1 mg QD.

In another embodiment, the dose is about 0.1 mg QD.

In another embodiment, the dose is less than 0.5 mg QD.

In another embodiment, the dose is about 0.5 mg QD.

In another embodiment, the dose is between about 0.1 and about 1.0 mgQD.

In another embodiment, the dose is between about 0.5 and about 1.0 mgQD.

In another embodiment, the dose is about 1 mg QD.

In another embodiment, the dose is about 2 mg QD.

In another embodiment, the dose is less than 0.2 mg QD.

In another embodiment, the dose is about 0.2 mg QD.

In another embodiment, the dose is less than 1.0 mg QD.

In another embodiment, the dose is about 1.0 mg QD.

In another embodiment, the dose is between about 0.2 and about 2.0 mgQD.

In another embodiment, the dose is between about 1.0 and about 2.0 mgQD.

In another embodiment, the dose is about 2 mg QD.

In another embodiment, the dose is about 4 mg QD.

In one embodiment, the amount of the compound administered is sufficientto provide a lung concentration of the compound at steady state, rangingfrom about 0.005 to about 100 μM, from about 0.005 to about 10 μM, fromabout 0.01 to about 10 μM, from about 0.01 to about 5 μM, from about0.005 to about 1 μM, from about 0.005 to about 0.5 μM, from about 0.005to about 0.5 μM, from about 0.01 to about 0.2 μM, or from about 0.01 toabout 0.1 μM. In one embodiment, the amount of the compound administeredis sufficient to provide a lung concentration at steady state, of about0.005 to about 100 μM. In another embodiment, the amount of the compoundadministered is sufficient to provide a lung concentration at steadystate, of about 0.005 to about 10 μM. In yet another embodiment, theamount of the compound administered is sufficient to provide a lungconcentration at steady state, of about 0.01 to about 10 μM. In yetanother embodiment, the amount of the compound administered issufficient to provide a lung concentration at steady state, of about0.01 to about 5 μM. In yet another embodiment, the amount of thecompound administered is sufficient to provide a lung concentration atsteady state, of about 0.005 to about 1 μM. In yet another embodiment,the amount of the compound administered is sufficient to provide a lungconcentration at steady state, of about 0.005 to about 0.5 μM. In yetanother embodiment, the amount of the compound administered issufficient to provide a lung concentration of the compound at steadystate, of about 0.01 to about 0.2 μM. In still another embodiment, theamount of the compound administered is sufficient to provide a lungconcentration of the compound at steady state, of about 0.01 to about0.1 μM. As used herein, the term “lung concentration at steady state” isthe concentration reached after a period of administration of acompound. Once steady state is reached, there are minor peaks andtroughs on the time dependent curve of the lung concentration of thecompound.

In one embodiment, the amount administered is sufficient to provide amaximum lung concentration (peak concentration) of the compound, rangingfrom about 0.005 to about 100 μM, from about 0.005 to about 10 μM, fromabout 0.01 to about 10 μM, from about 0.01 to about 5 μM, from about0.005 to about 1 μM, from about 0.005 to about 0.5 μM, from about 0.01to about 0.2 μM, or from about 0.01 to about 0.1 μM. In one embodiment,the amount of the compound administered is sufficient to provide amaximum lung concentration of the compound of about 0.005 to about 100μM. In another embodiment, the amount of the compound administered issufficient to provide a maximum lung concentration of the compound ofabout 0.005 to about 10 μM. In yet another embodiment, the amount of thecompound administered is sufficient to provide a maximum lungconcentration of the compound of about 0.01 to about 10 μM. In yetanother embodiment, the amount of the compound administered issufficient to provide a maximum lung concentration of the compound ofabout 0.01 to about 5 μM. In yet another embodiment, the amount of thecompound administered is sufficient to provide a maximum lungconcentration of the compound of about 0.005 to about 1 μM. In yetanother embodiment, the amount of the compound administered issufficient to provide a maximum lung concentration of the compound ofabout 0.005 to about 0.5 μM. In yet another embodiment, the amount ofthe compound administered is sufficient to provide a maximum lungconcentration of the compound of about 0.01 to about 0.2 μM. In stillanother embodiment, the amount of the compound administered issufficient to provide a maximum lung concentration of the compound ofabout 0.01 to about 0.1 μM.

In one embodiment, the amount administered is sufficient to provide aminimum lung concentration (trough concentration) of the compound,ranging from about 0.005 to about 100 μM, from about 0.005 to about 10μM, from about 0.01 to about 10 μM, from about 0.01 to about 5 μM, fromabout 0.005 to about 1 μM, about 0.005 to about 0.5 μM, from about 0.01to about 0.2 μM, or from about 0.01 to about 0.1 μM, when more than onedoses are administered. In one embodiment, the amount of the compoundadministered is sufficient to provide a minimum lung concentration ofthe compound of about 0.005 to about 100 μM. In another embodiment, theamount of the compound administered is sufficient to provide a minimumlung concentration of the compound of about 0.005 to about 10 μM. In yetanother embodiment, the amount of the compound administered issufficient to provide a minimum lung concentration of the compound ofabout 0.01 to about 10 μM. In yet another embodiment, the amount of thecompound administered is sufficient to provide a minimum lungconcentration of the compound of about 0.01 to about 5 μM. In yetanother embodiment, the amount of the compound administered issufficient to provide a minimum lung concentration of the compound ofabout 0.005 to about 1 μM. In yet another embodiment, the amount of thecompound administered is sufficient to provide a minimum lungconcentration of the compound of about 0.005 to about 0.5 μM. In yetanother embodiment, the amount of the compound administered issufficient to provide a minimum lung concentration of the compound ofabout 0.01 to about 0.2 μM. In still another embodiment, the amount ofthe compound administered is sufficient to provide a minimum lungconcentration of the compound of about 0.01 to about 0.1 μM.

In one embodiment, the amount administered is sufficient to provide anarea under the curve (AUC) of the compound, ranging from about 50 toabout 10,000 ng*hr/mL, about 100 to about 50,000 ng*hr/mL, from about100 to 25,000 ng*hr/mL, or from about 10,000 to 25,000 ng*hr/mL. Incertain embodiments, a compound provided herein (e.g., Compound 4), oran enantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof, isadministered once per day from one day to six months, from one week tothree months, from one week to four weeks, from one week to three weeks,or from one week to two weeks.

In certain embodiments, the compound provided herein is administeredonce per day for one week, two weeks, three weeks, or four weeks. In oneembodiment, the compound provided herein is administered once per dayfor one week. In another embodiment, the compound provided herein isadministered once per day for two weeks. In yet another embodiment, thecompound provided herein is administered once per day for three weeks.In still another embodiment, the compound provided herein isadministered once per day for four weeks.

In certain embodiments, a compound provided herein (e.g., Compound 4),or an enantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof, isadministered twice per day from one day to six months, from one week tothree months, from one week to four weeks, from one week to three weeks,or from one week to two weeks. In certain embodiments, the compoundprovided herein is administered twice per day for one week, two weeks,three weeks, or four weeks. In one embodiment, the compound providedherein is administered twice per day for one week. In anotherembodiment, the compound provided herein is administered twice per dayfor two weeks. In yet another embodiment, the compound provided hereinis administered twice per day for three weeks. In still anotherembodiment, the compound provided herein is administered twice per dayfor four weeks.

The compound provided herein (e.g., Compound 4), or an enantiomer, amixture of enantiomers, or a mixture of two or more diastereomersthereof, or a pharmaceutically acceptable form thereof, can be deliveredas a single dose such as, e.g., a single bolus injection, or oraltablets or pills; or over time, such as, e.g., continuous infusion overtime or divided bolus doses over time. The compound can be administeredrepeatedly if necessary, for example, until the patient experiencesstable disease or regression, or until the patient experiences diseaseprogression or unacceptable toxicity.

Chronic Obstructive Pulmonary Disease

In one embodiment, said obstructive lung disease or disorder is chronicobstructive pulmonary disease (COPD), e.g., as diagnosed by a forcedexpiratory air volume in 1 second (FEV₁) to forced vital capacity (FVC)ratio of less than 0.7. In another embodiment, administration of acompound provided herein results in a detectable rise in the FEV₁/FECratio above 0.7 after administration, e.g., a rise of 0.02, 0.03, 0.04,0.05, 0.06, 0.07, 0.08, 0.09, 0.10, or more or more.

In one embodiment, provided herein is a method of reducing a COPDassociated symptom in a subject, comprising administering a compoundprovided herein (e.g., Compound 4), or an enantiomer, a mixture ofenantiomers, or a mixture of two or more diastereomers thereof, or apharmaceutically acceptable form thereof, by inhalation in an amountsufficient to reduce the COPD associated symptom. In one embodiment, thesubject is a mammalian subject, e.g., an animal model or as part oftherapeutic protocol. In one embodiment, the compound is used as asingle agent or in combination with another agent or therapeuticmodality.

In one embodiment, provided herein is a method of treating, preventing,and/or managing COPD in a subject, comprising administering an effectiveamount of a compound provided herein (e.g., Compound 4), or anenantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof, toa subject in need thereof by inhalation. In one embodiment, the compoundis administered as a single agent. In another embodiment, the compoundis administered in combination with another agent or therapeuticmodality.

As used herein, and unless otherwise specified, “COPD” or a “symptom”associated with COPD encompasses all types of manifestation of COPD asdisclosed herein or as known in the art. Examples of COPD include, butare not limited to, emphysema, chronic bronchitis, and bronchiectasis.Examples of symptom of COPD include, but are not limited to, wheezing,coughing, chest tightness, shortness of breath, difficulty in breathing,coughing up mucus/phlegm, and use of accessory muscle. Symptoms areoften worse at night or in the early morning, or in response to exerciseor cold air. In one embodiment, the symptom of asthma is shortness ofbreath or difficulty in breathing.

As used herein, and unless otherwise specified, to “decrease,”“ameliorate,” “reduce,” “inhibit,” “treat” (or the like) COPD or asymptom associated with COPD includes reducing the severity and/orfrequency of one or more symptoms of COPD, as well as preventing COPDand/or one or more symptoms of COPD (e.g., by reducing the severityand/or frequency of flares of symptoms).

In some embodiments, the symptom is reduced by at least about 2%, atleast about 5%, at least about 10%, at least about 15%, at least about20%, at least about 25%, at least about 30%, at least about 40%, atleast about 50%, at least about 60%, at least about 70%, at least about80%, at least about 90%, or at least about 95% relative to a controllevel. The control level includes any appropriate control as known inthe art. For example, the control level can be the pre-treatment levelin the sample or subject treated, or it can be the level in a controlpopulation (e.g., the level in subjects who do not have COPD or thelevel in samples derived from subjects who do not have COPD). In someembodiments, the decrease is statistically significant, for example, asassessed using an appropriate parametric or non-parametric statisticalcomparison.

In certain embodiments, the subject is an animal model of COPD, a humanwith COPD, or a subject (e.g., a human) at risk for developing COPD. Insome embodiments, the subject is a human who has a family history ofCOPD, who carries a gene associated with COPD, who is positive for abiomarker associated with COPD, or a combination thereof. In someembodiments, the subject has been diagnosed with COPD. In someembodiments, the subject has one or more signs or symptoms associatedwith COPD. In some embodiments, the subject is at risk for developingCOPD (e.g., the subject carries a gene that, individually, or incombination with other genes or environmental factors, is associatedwith development of COPD).

In one embodiment, the subject has been previously diagnosed of COPD orhas episodic symptoms of airflow obstruction (e.g., shortness of breath,wheezing and/or chest tightness) for at least 1 week, 2 weeks, 1 month,2 months, 3 months, 6 months, 9 months, 12 months before a compoundprovided herein (e.g., Compound 4), or an enantiomer, a mixture ofenantiomers, or a mixture of two or more diastereomers thereof, or apharmaceutically acceptable form thereof f, is administered. In oneembodiment, the subject has been previously diagnosed of COPD or hasepisodic symptoms of airflow obstruction (e.g., wheezing and/or chesttightness) for at least 6 months before a compound provided herein(e.g., Compound 4), or an enantiomer or a mixture of enantiomersthereof, or a pharmaceutically acceptable salt, solvate, hydrate,co-crystal, clathrate, or polymorph thereof, is administered.

In some embodiments, the subject has been previously treated for COPD.In some embodiments, the subject has been previously treated for COPDbut are non-responsive to standard therapies. In one embodiment, thestandard therapy is steroid, e.g., corticosteroids. In some embodiments,the subject has developed steroid resistance, e.g., from previoustreatment with steroids. In some embodiments, the subject can haveinherent steroid resistance that is not a result of previous treatments.Steroid resistance can be overcome by a PI3K inhibitor, e.g., a compoundprovided herein (e.g., Compound 4). Thus, combination therapy with acompound provided herein and a steroid can be beneficial. In oneembodiment, provided herein is a method of treating, preventing, and/ormanaging COPD in a subject, comprising administering an effective amountof a compound provided herein (e.g., Compound 4), or an enantiomer, amixture of enantiomers, or a mixture of two or more diastereomersthereof, or a pharmaceutically acceptable form thereof, to a subject inneed thereof, wherein the subject has been previously administered atherapy for COPD.

In some embodiments, the subject has not been previously treated forCOPD.

In one embodiment, without being limited by any particular theory,administering an effective amount of a compound provided herein (e.g.,Compound 4), or an enantiomer, a mixture of enantiomers, or a mixture oftwo or more diastereomers thereof, or a pharmaceutically acceptable formthereof, does not result in, or results in reduced, one or more commonside effects of COPD treatment. The common side effects of COPDtreatment include, but are not limited to: allergic reactions such asrashes, hives, swelling of the face, mouth and tongue, and breathingproblems; sudden breathing problems; effects on heart such as increasedblood pressure, fast and irregular heart beat, and chest pain; effectson nervous system such as tremor and nervousness; reduced adrenalfunction; changes in blood contents; weakened immune system and higherchance of infections; lower bone mineral density; eye problems such asglaucoma and cataracts; slowed growth in children; pneumonia; thrush inthe mouth and throat; throat irritation; hoarseness and voice changes;viral respiratory infections; headache; and muscle and bone pain.

In some embodiments, the side effect is reduced by at least about 2%, atleast about 5%, at least about 10%, at least about 15%, at least about20%, at least about 25%, at least about 30%, at least about 40%, atleast about 50%, at least about 60%, at least about 70%, at least about80%, at least about 90%, or at least about 95% relative to a controllevel. The control level includes any appropriate control as known inthe art. For example, the control level can be the side effect level inthe subject treated with other COPD therapies (e.g., albuterol,levalbuterol, ipratropium, tiotropium, terbutaline, theophylline,formoterol, salmeterol, flucatisone, methylprednisone, and prednisone).In some embodiments, the decrease is statistically significant, forexample, as assessed using an appropriate parametric or non-parametricstatistical comparison.

Asthma

In another specific embodiment, said obstructive lung disease ordisorder is asthma. In some embodiments, administration of a compoundprovided herein results in a detectable improvement in one or moresymptoms of asthma, e.g., airway obstruction, as determined byspirometry or a peak flow meter.

In one embodiment, provided herein is a method of reducing an asthmaassociated symptom in a subject, comprising administering a compoundprovided herein (e.g., Compound 4), or an enantiomer, a mixture ofenantiomers, or a mixture of two or more diastereomers thereof, or apharmaceutically acceptable form thereof, by inhalation in an amountsufficient to reduce the asthma associated symptom. In one embodiment,the subject is a mammalian subject, e.g., an animal model or as part oftherapeutic protocol. In one embodiment, the compound is used as asingle agent or in combination with another agent or therapeuticmodality.

In one embodiment, provided herein is a method of treating, preventing,and/or managing asthma in a subject, comprising administering aneffective amount of a compound provided herein (e.g., Compound 4), or anenantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof toa subject in need thereof by inhalation. In one embodiment, the compoundis administered as a single agent. In another embodiment, the compoundis administered in combination with another agent or therapeuticmodality.

As used herein, and unless otherwise specified, “asthma” or a “symptom”associated with asthma encompasses all types of manifestation of asthmaas disclosed herein or as known in the art. Examples of asthma include,but are not limited to, severe and/or refractory asthma, atopic(extrinsic) asthma, non-atopic (intrinsic) asthma, type 1 brittleasthma, type 2 brittle asthma, asthma attack, status asthmaticus,exercise-induced asthma, or occupational asthma. In one embodiment, theasthma is severe or refractory asthma. Examples of symptom of asthmainclude, but are not limited to, wheezing, coughing, chest tightness,shortness of breath, and use of accessory muscle. Symptoms are oftenworse at night or in the early morning, or in response to exercise orcold air. Asthma is clinically classified according to the frequency ofsymptoms, forced expiratory volume in 1 second (FEV₁), and peakexpiratory flow rate. In one embodiment, the symptom of asthma iswheezing or chest tightness.

As used herein, and unless otherwise specified, “asthma” or a “symptom”associated with asthma also encompasses biological concomitants ofasthma as disclosed herein or as known in the art. Examples include, butare not limited to, immune complexes, elevated levels of cytokines(e.g., interferons (e.g., Type I interferons, e.g., IFN-α and/or IFN-β);interleukins (e.g., IL-6, IL-8, IL-1, and IL-18) and TNF-α), elevatedlevels of anti-dsDNA autoantibodies, overexpression of IFN-α and/orIFN-β inducible genes, elevated levels of IP-10, elevated levels ofsCD40L, reduced levels of C3-derived C3b, reduced peripheral iNKT cellfrequencies, defective B cell-mediated stimulation of iNKT cells,altered CD1d expression on B cells, reduced numbers of naturalregulatory T cells (Treg), altered level of C-reactive protein,overexpression of mRNA for IL-4, overexpression of mRNA for IL-21, andelevated serium anti-collagen level. In some embodiments, the symptom isoverexpression of IFN-α, TNF-α, IL-δ, IL-8, or IL-1. In one embodiment,the symptom is overexpression of IFN-α. In one embodiment, the symptomis overexpression of IL-6. In some embodiments, the symptom isoverexpression of mRNA for IL-4 or overexpression of mRNA for IL-21. Insome embodiments, the symptom is elevated serium anti-collagen level.

As used herein, and unless otherwise specified, to “decrease,”“ameliorate,” “reduce,” “inhibit,” “treat” (or the like) asthma or asymptom associated with asthma includes reducing the severity and/orfrequency of one or more symptoms of asthma, as well as preventingasthma and/or one or more symptoms of asthma (e.g., by reducing theseverity and/or frequency of flares of symptoms).

In some embodiments, the symptom is reduced by at least about 2%, atleast about 5%, at least about 10%, at least about 15%, at least about20%, at least about 25%, at least about 30%, at least about 40%, atleast about 50%, at least about 60%, at least about 70%, at least about80%, at least about 90%, or at least about 95% relative to a controllevel. The control level includes any appropriate control as known inthe art. For example, the control level can be the pre-treatment levelin the sample or subject treated, or it can be the level in a controlpopulation (e.g., the level in subjects who do not have asthma or thelevel in samples derived from subjects who do not have asthma). In someembodiments, the decrease is statistically significant, for example, asassessed using an appropriate parametric or non-parametric statisticalcomparison.

In certain embodiments, the subject is an animal model of asthma, ahuman with asthma, or a subject (e.g., a human) at risk for developingasthma. In some embodiments, the subject is a human who has a familyhistory of asthma, who carries a gene associated with asthma, who ispositive for a biomarker associated with asthma, or a combinationthereof. In some embodiments, the subject has been diagnosed withasthma. In some embodiments, the subject has one or more signs orsymptoms associated with asthma. In some embodiments, the subject is atrisk for developing asthma (e.g., the subject carries a gene that,individually, or in combination with other genes or environmentalfactors, is associated with development of asthma).

In one embodiment, the subject has been previously diagnosed of asthmaor has episodic symptoms of airflow obstruction (e.g., wheezing and/orchest tightness) for at least 1 week, 2 weeks, 1 month, 2 months, 3months, 6 months, 9 months, 12 months before a compound provided herein(e.g., Compound 4), or an enantiomer, a mixture of enantiomers, or amixture of two or more diastereomers thereof, or a pharmaceuticallyacceptable form thereof, is administered. In one embodiment, the subjecthas been previously diagnosed of asthma or has episodic symptoms ofairflow obstruction (e.g., wheezing and/or chest tightness) for at least6 months before a compound provided herein (e.g., Compound 4), or anenantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof, isadministered.

In one embodiment, the subject has a forced expiratory volume in onesecond (FEV₁) value of at least 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%,55%, or 50% of a control value. In one embodiment, the subject has aforced expiratory volume in one second (FEV₁) value of at least 70% of acontrol value. In one embodiment, the control value may be calculatedbased on American Thoracic Society (ATS)/European Respiratory Society(ERS) standards.

In one embodiment, the subject has a positive response to a skin pricktest to an allergen. In one embodiment, the positive response means thatthe induration of skin test wheal is larger in diameter (e.g., at least2 mm larger) than the diameter of the control wheal. The allergen can beany allergen provided herein or known in the art that can be used in thediagnosis or determining status of asthma.

In one embodiment, the subject has an early-phase asthmatic response(EAR) of at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, or 50% toan inhaled allergen challenge. In one embodiment, the subject has anearly-phase asthmatic response of at least 20% to an inhaled allergenchallenge. In one embodiment, the EAR response is a decrease frompre-challenge in FEV₁ on 2 consecutive occasions within 0 to <3 hours oflast allergen challenge.

In one embodiment, the subject has a late-phase asthmatic response (LAR)of at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, or 50% to aninhaled allergen challenge. In one embodiment, the subject has alate-phase asthmatic response of at least 15% to an inhaled allergenchallenge. In one embodiment, the LAR response is a decrease frompre-challenge in FEV₁ on 2 consecutive occasions within 3 to 10 hours oflast allergen challenge.

In one embodiment, the subject has an early-phase asthmatic response ofat least 20% and a late-phase asthmatic response of at least 15% to aninhaled allergen challenge. The inhaled allergen can be any inhaledallergen provided herein or known in the art that can be used in thediagnosis or determining status of asthma.

In one embodiment, the subject exhibits an elevated level of C-reactiveprotein. In one embodiment, the subject exhibits an elevated level ofC-reactive protein of at least 1.0 mg/L. In one embodiment, the subjectexhibits an elevated level of C-reactive protein of at least 7 mg/L.

In some embodiments, the subject exhibits elevated levels of antinuclearantibodies (e.g., anti-Smith antibodies, anti-double stranded DNA(dsDNA) antibodies, anti-U1 RNP, SS-a (or anti-Ro), SS-b (or anti-La)),antiphospholipid antibodies, anti-ss DNA antibodies, anti-histoneantibodies, or anticardiolipin antibodies. In some embodiments, thesubject exhibits elevated levels of anti-dsDNA antibodies. In someembodiments, the subject exhibits elevated levels of anti-Sm antibodies.

In some embodiments, the subject exhibits autoantibodies against one ormore antigens that are known to be associated with asthma or with asthmasubtypes. In some embodiments, the subject exhibits autoantibodiesagainst Sm/anti-RNP or Ro/La autoantigens.

The levels of antibodies associated with asthma can be assessed usingany suitable method, e.g., methods known in the art, e.g., indirectimmunofluorescence. In some embodiments, the methods disclosed hereinreduce or prevent an increase in the levels of one or more of theforegoing antibodies.

In some embodiments, the subject exhibits elevated levels of IFN-α,TNF-α, IL-6, IL-8, or IL-1. In one embodiment, the subject exhibits anelevated level of IFN-α. In another embodiment, the subject exhibits anelevated level of IL-6. In another embodiment, the subject exhibits anelevated level of mRNA for IL-4 or IL-21.

In some embodiments, the subject has a mutation (e.g., an SNP) in a geneassociated with asthma. In one embodiment, the gene is selected fromSTAT4, IRF5, BANK1, ITGAM, PD1, FAM167A-BLK, IRF5-TNP03, KIAA1542,TNFAIP3, XKR6, 1q25.1, PXK, ATG5, ICA1, XKR6, LYN and SCUB2 or acombination thereof. In some embodiments, the subject carries the DR3and DQ2 variants, or the DR2 and DQ6 variants of HLA class II genes. Insome embodiments, the subject has a deficiency in one or more complementproteins, e.g. a deficiency of a complement protein coded by the C4A orC2 genes on chromosome 6, or the C1r and C1s genes on chromosome 12.

In some embodiments, the subject exhibits excessive PI3K activity orabnormal activity (e.g., excessive or reduced activity) of one or morecomponents of the PI3K signaling pathway (e.g., Akt (PKB), mTOR, a Teckinase (e.g., Btk, Itk, Tec), phospholipase C, PDK1, PKCs, NFκB, Rac GEF(e.g., Vav-1), or Rac).

In some embodiments, the subject is an animal model of asthma providedherein or known in the art. Examples include, but are not limited to,the murine lipopolysaccharide (LPS) induced pulmonary inflammationmodel, and the murine ovalbumin-induced allergic airway inflammationmodel.

In some embodiments, the subject has been previously treated for asthma.In some embodiments, the subject has been previously treated for asthmabut are non-responsive to standard therapies. Thus, in one embodiment,provided herein is a method of treating, preventing, and/or managingasthma in a subject, comprising administering an effective amount of acompound provided herein (e.g., Compound 4), or an enantiomer, a mixtureof enantiomers, or a mixture of two or more diastereomers thereof, or apharmaceutically acceptable form thereof, to a subject in need thereof,wherein the subject has been previously administered a therapy forasthma.

In some embodiments, the subject has not been previously treated forasthma.

In one embodiment, without being limited by any particular theory,administering an effective amount of a compound provided herein (e.g.,Compound 4), or an enantiomer, a mixture of enantiomers, or a mixture oftwo or more diastereomers thereof, or a pharmaceutically acceptable formthereof, does not result in, or results in reduced, one or more commonside effects of asthma treatment. The common side effects of asthmatreatment include, but are not limited to, oral candidiasis, thrush,dysphonia (hoarseness), reflex cough, bronchospasm, poor growth,decreased bone density, disseminated varicella infection (chickenpoxthat spreads to organs), easy bruising, cataracts, glaucoma, adrenalgland suppression, stomach upset, headache, liver test abnormalities,skin rashes, Churg Strauss syndrome, bad taste in month, cough, itching,sore throat, sneezing, stuffy nose, shortness of breath, wheezing, viralillness, upper respiratory tract infections, sinusitis, feeling dizzy orfaint, hives, changes in voice, swelling of the tongue, or difficulty inswallowing.

In some embodiments, the side effect is reduced by at least about 2%, atleast about 5%, at least about 10%, at least about 15%, at least about20%, at least about 25%, at least about 30%, at least about 40%, atleast about 50%, at least about 60%, at least about 70%, at least about80%, at least about 90%, or at least about 95% relative to a controllevel. The control level includes any appropriate control as known inthe art. For example, the control level can be the side effect level inthe subject treated with other asthma therapies (e.g., Xolair, CromolynSodium, Nedocromil, Montelukast, and prednisone). In some embodiments,the decrease is statistically significant, for example, as assessedusing an appropriate parametric or non-parametric statisticalcomparison.

In one embodiment, the regression of asthma is a decrease (e.g., atleast 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50% decrease) in thelevel of maximal decrease from pre-allergen challenge in FEV₁ followingallergen challenge. The level of maximal decrease from pre-allergenchallenge in FEV₁ following allergen challenge can be measured in EAR orLAR.

In one embodiment, the regression of asthma is a decrease (e.g., atleast 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50% decrease) in areaunder the curve (AUC) of FEV₁ following allergen challenge.

In one embodiment, the regression of asthma is an increase (e.g., atleast 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50% increase) in theamount of methacholine that is required to induce a 20% fall in FEV₁(PC₂₀) following allergen challenge.

In one embodiment, the regression of asthma is a decrease (e.g., atleast 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50% decrease) inexhaled nitric oxide level of the subject.

In one embodiment, the regression of asthma is a decrease (e.g., atleast 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50% decrease) in theC-reactive protein (CRP) level of the subject.

In one embodiment, the regression of asthma is a decrease (e.g., atleast 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50% decrease) in whiteblood cell count and/or differential cell count in induced sputum of thesubject after allergen challenge.

Combination Therapy

In some embodiments, provided herein are methods for combinationtherapies in which an agent known to modulate other pathways, or othercomponents of the same pathway, or even overlapping sets of targetenzymes are used in combination with a compound provided herein, or apharmaceutically acceptable form (e.g., pharmaceutically acceptablesalts, hydrates, solvates, isomers, prodrugs, and isotopically labeledderivatives) thereof. In one aspect, such therapy includes, but is notlimited to, the combination of the subject compound withchemotherapeutic agents, therapeutic antibodies, and radiationtreatment, to provide a synergistic or additive therapeutic effect.

By “in combination with,” it is not intended to imply that the othertherapy and the PI3K modulator must be administered at the same timeand/or formulated for delivery together, although these methods ofdelivery are within the scope of this disclosure. The compound providedherein can be administered concurrently with, prior to (e.g., 5 minutes,15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours,12 hours, 24 hours, 48 hours, 72 hours, 96 hours, 1 week, 2 weeks, 3weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, 12 weeks, or 16 weeksbefore), or subsequent to (e.g., 5 minutes, 15 minutes, 30 minutes, 45minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6weeks, 8 weeks, 12 weeks, or 16 weeks after), one or more othertherapies (e.g., one or more other additional agents). In general, eachtherapeutic agent will be administered at a dose and/or on a timeschedule determined for that particular agent. The other therapeuticagent can be administered with the compound provided herein in a singlecomposition or separately in a different composition. Triple therapy isalso contemplated herein.

In general, it is expected that additional therapeutic agents employedin combination be utilized at levels that do not exceed the levels atwhich they are utilized individually. In some embodiments, the levelsutilized in combination will be lower than those utilized individually.

In some embodiments, the compound provided herein is a first linetreatment for cancer or hematologic malignancy, i.e., it is used in asubject who has not been previously administered another drug or therapyintended to treat cancer or hematologic malignancy or one or moresymptoms thereof.

In other embodiments, the compound provided herein is a second linetreatment for cancer or hematologic malignancy, i.e., it is used in asubject who has been previously administered another drug or therapyintended to treat cancer or hematologic malignancy or one or moresymptoms thereof.

In other embodiments, the compound provided herein is a third or fourthline treatment for cancer or hematologic malignancy, i.e., it is used ina subject who has been previously administered two or three other drugsor therapies intended to treat cancer or hematologic malignancy or oneor more symptoms thereof.

In embodiments where two agents are administered, the agents can beadministered in any order. For example, the two agents can beadministered concurrently (i.e., essentially at the same time, or withinthe same treatment) or sequentially (i.e., one immediately following theother, or alternatively, with a gap in between administration of thetwo). In some embodiments, the compound provided herein is administeredsequentially (i.e., after the first therapeutic).

In one aspect, a compound provided herein, or a pharmaceuticallyacceptable form (e.g., pharmaceutically acceptable salts, hydrates,solvates, isomers, prodrugs, and isotopically labeled derivatives)thereof, or a pharmaceutical composition as provided herein, can presentsynergistic or additive efficacy when administered in combination withagents that inhibit IgE production or activity. Such combination canreduce the undesired effect of high level of IgE associated with the useof one or more PI3K-δ inhibitors, if such effect occurs. This can beparticularly useful in treatment of autoimmune and inflammatorydisorders (AIID) such as rheumatoid arthritis. Additionally, theadministration of PI3K-δ, PI3K-γ, or PI3K-δ/γ inhibitors as providedherein in combination with inhibitors of mTOR can also exhibit synergythrough enhanced inhibition of the PI3K pathway.

In a separate but related aspect, provided herein is a combinationtreatment of a disease associated with PI3K-δ comprising administeringto a subject in need thereof a PI3K-δ inhibitor and an agent thatinhibits IgE production or activity. Other exemplary PI3K-δ inhibitorsare applicable for this combination and they are described in, e.g.,U.S. Pat. No. 6,800,620, incorporated herein by reference. Suchcombination treatment is particularly useful for treating autoimmune andinflammatory diseases (AIID) including, but not limited to rheumatoidarthritis.

Agents that inhibit IgE production are known in the art and theyinclude, but are not limited to, one or more of TEI-9874,2-(4-(6-cyclohexyloxy-2-naphtyloxy)phenylacetamide)benzoic acid,rapamycin, rapamycin analogs (i.e., rapalogs), TORC1 inhibitors, TORC2inhibitors, and any other compounds that inhibit mTORC1 and mTORC2.Agents that inhibit IgE activity include, for example, anti-IgEantibodies such as for example Omalizumab and TNX-901.

For treatment of autoimmune diseases, a compound provided herein, or apharmaceutically acceptable form (e.g., pharmaceutically acceptablesalts, hydrates, solvates, isomers, prodrugs, and isotopically labeledderivatives) thereof, or a pharmaceutical composition as providedherein, can be used in combination with commonly prescribed drugsincluding, but not limited to, Enbrel®, Remicade®, Humira®, Avonex®, andRebif®. For treatment of respiratory diseases, the subject compounds, orpharmaceutically acceptable forms thereof, or pharmaceuticalcompositions, can be administered in combination with commonlyprescribed drugs including, but not limited to, Xolair®, Advair®,Singulair®, and Spiriva®.

The compounds as provided herein, or pharmaceutically acceptable forms(e.g., pharmaceutically acceptable salts, hydrates, solvates, isomers,prodrugs, and isotopically labeled derivatives) thereof, orpharmaceutical compositions as provided herein, can be formulated oradministered in conjunction with other agents that act to relieve thesymptoms of inflammatory conditions such as encephalomyelitis, asthma,and the other diseases described herein. These agents includenon-steroidal anti-inflammatory drugs (NSAIDs), e.g., acetylsalicylicacid; ibuprofen; naproxen; indomethacin; nabumetone; tolmetin; etc.Corticosteroids are used to reduce inflammation and suppress activity ofthe immune system. An exemplary drug of this type is Prednisone.Chloroquine (Aralen) or hydroxychloroquine (Plaquenil) can also be usedin some individuals with lupus. They can be prescribed for skin andjoint symptoms of lupus. Azathioprine (Imuran) and cyclophosphamide(Cytoxan) suppress inflammation and tend to suppress the immune system.Other agents, e.g., methotrexate and cyclosporin are used to control thesymptoms of lupus. Anticoagulants are employed to prevent blood fromclotting rapidly. They range from aspirin at very low dose whichprevents platelets from sticking, to heparin/coumadin. Other compoundsused in the treatment of lupus include belimumab (Benlysta®).

In another aspect, provided herein is a pharmaceutical composition forinhibiting abnormal cell growth in a subject which comprises an amountof a compound provided herein, or a pharmaceutically acceptable form(e.g., pharmaceutically acceptable salts, hydrates, solvates, isomers,prodrugs, and isotopically labeled derivatives) thereof, in combinationwith an amount of an anti-cancer agent (e.g., a chemotherapeutic agentor a biotherapeutic agent). Many chemotherapeutics are presently knownin the art and can be used in combination with a compound providedherein.

In some embodiments, the chemotherapeutic is selected from mitoticinhibitors, alkylating agents, anti-metabolites, intercalatingantibiotics, growth factor inhibitors, cell cycle inhibitors, enzymes,topoisomerase inhibitors, biological response modifiers, anti-hormones,angiogenesis inhibitors, and anti-androgens. Non-limiting examples arechemotherapeutic agents, cytotoxic agents, and non-peptide smallmolecules such as Gleevec® (imatinib mesylate), Velcade® (bortezomib),Casodex™ (bicalutamide), Iressa® (gefitinib), Tarceva® (erlotinib), andAdriamycin® (doxorubicin) as well as a host of chemotherapeutic agents.Non-limiting examples of chemotherapeutic agents include alkylatingagents such as thiotepa and cyclosphosphamide (CYTOXAN™); alkylsulfonates such as busulfan, improsulfan and piposulfan; aziridines suchas benzodopa, carboquone, meturedopa, and uredopa; ethylenimines andmethylamelamines including altretamine, triethylenemelamine,trietylenephosphoramide, triethylenethiophosphoramide andtrimethylolomelamine; BTK inhibitors such as ibrutinib (PCI-32765),AVL-292, Dasatinib, LFM-AI3, ONO-WG-307, and GDC-0834; HDAC inhibitorssuch as vorinostat, romidepsin, panobinostat, valproic acid, belinostat,mocetinostat, abrexinostat, entinostat, SB939, resminostat, givinostat,CUDC-101, AR-42, CHR-2845, CHR-3996, 4SC-202, CG200745, ACY-1215 andkevetrin; EZH2 inhibitors such as, but not limited to, EPZ-6438(N-((4,6-dimethyl-2-oxo-1,2-dihydropyridin-3-yl)methyl)-5-(ethyl(tetrahydro-2H-pyran-4-yl)amino)-4-methyl-4′-(morpholinomethyl)-[1,1′-biphenyl]-3-carboxamide),GSK-126((S)-1-(sec-butyl)-N-((4,6-dimethyl-2-oxo-1,2-dihydropyridin-3-yl)methyl)-3-methyl-6-(6-(piperazin-1-yl)pyridin-3-yl)-1H-indole-4-carboxamide),GSK-343(1-Isopropyl-N-((6-methyl-2-oxo-4-propyl-1,2-dihydropyridin-3-yl)methyl)-6-(2-(4-methylpiperazin-1-yl)pyridine-4-yl)-1H-indazole-4-carboxamide),E11, 3-deazaneplanocin A (DNNep,5R-(4-amino-1H-imidazo[4,5-c]pyridin-1-yl)-3-(hydroxymethyl)-3-cyclopentene-1S,2R-diol),small interfering RNA (siRNA) duplexes targeted against EZH2 (S. M.Elbashir et al., Nature 411:494-498 (2001)), isoliquiritigenin, andthose provided in, for example, U.S. Publication Nos. 2009/0012031,2009/0203010, 2010/0222420, 2011/0251216, 2011/0286990, 2012/0014962,2012/0071418, 2013/0040906, and 2013/0195843, all of which areincorporated herein by reference; JAK/STAT inhibitors such aslestaurtinib, tofacitinib, ruxolitinib, pacritinib, CYT387, baricitinib,GLPG0636, TG101348, INCB16562, CP-690550, and AZD1480; PKC-β inhibitorsuch as Enzastaurin; SYK inhibitors such as, but not limited to,GS-9973, PRT 062607, R406, (S)-2-(2-((3,5-dimethylphenyl)amino)pyrimidin-4-yl)-N-(1-hydroxypropan-2-yl)-4-methylthiazole-5-carboxamide,R112, GSK143, BAY61-3606, PP2, PRT 060318, R348, and those provided in,for example, U.S. Publication Nos. 2003/0113828, 2003/0158195,2003/0229090, 2005/0075306, 2005/0232969, 2005/0267059, 2006/0205731,2006/0247262, 2007/0219152, 2007/0219195, 2008/0114024, 2009/0171089,2009/0306214, 2010/0048567, 2010/0152159, 2010/0152182, 2010/0316649,2011/0053897, 2011/0112098, 2011/0245205, 2011/0275655, 2012/0027834,2012/0093913, 2012/0101275, 2012/0130073, 2012/0142671, 2012/0184526,2012/0220582, 2012/0277192, 2012/0309735, 2013/0040984, 2013/0090309,2013/0116260, and 2013/0165431, all of which are incorporated herein byreference; SYK inhibitor such as R788 (fostamatinib); SYK/JAK dualinhibitor such as PRT2070; nitrogen mustards such as bendamustine,chlorambucil, chlornaphazine, cholophosphamide, estramustine,ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride,melphalan, novembichin, phenesterine, prednimustine, trofosfamide,uracil mustard; nitrosureas such as carmustine, chlorozotocin,fotemustine, lomustine, nimustine, ranimustine; antibiotics such asaclacinomycins, actinomycin, authramycin, azaserine, bleomycins,cactinomycin, calicheamicin, carabicin, carminomycin, carzinophilin,chromomycins, dactinomycin, daunorubicin, detorubicin,6-diazo-5-oxo-L-norleucine, doxorubicin, epirubicin, esorubicin,idarubicin, marcellomycin, mitomycin C, mycophenolic acid, nogalamycin,olivomycins, peplomycin, porfiromycin, puromycin, quelamycin,rodorubicin, streptonigrin, streptozocin, tubercidin, ubenimex,zinostatin, zorubicin; anti-metabolites such as methotrexate and5-fluorouracil (5-FU); folic acid analogues such as denopterin,methotrexate, pralatrexate, pteropterin, trimetrexate; purine analogssuch as fludarabine, 6-mercaptopurine, thiamiprine, thioguanine;pyrimidine analogs such as ancitabine, azacitidine, 6-azauridine,carmofur, cytarabine, dideoxyuridine, doxifluridine, enocitabine,floxuridine, androgens such as calusterone, dromostanolone propionate,epitiostanol, mepitiostane, testolactone; anti-adrenals such asaminoglutethimide, mitotane, trilostane; folic acid replenisher such asfolinic acid; aceglatone; aldophosphamide glycoside; aminolevulinicacid; amsacrine; bestrabucil; bisantrene; edatrexate; defofamine;demecolcine; diaziquone; elfomithine; elliptinium acetate; etoglucid;gallium nitrate; hydroxyurea; lentinan; lonidamine; mitoguazone;mitoxantrone; mopidamol; nitracrine; pentostatin; phenamet; pirarubicin;podophyllinic acid; 2-ethylhydrazide; procarbazine; PSK.R™; razoxane;sizofiran; spirogermanium; tenuazonic acid; triaziquone;2,2′,2″-trichlorotriethyla-mine; urethan; vindesine; dacarbazine;mannomustine; mitobronitol; mitolactol; pipobroman; gacytosine;arabinoside (Ara-C); cyclophosphamide; thiotepa; taxanes, e.g.,paclitaxel (e.g., TAXOL™) and docetaxel (e.g., TAXOTERE™) and ABRAXANE®(paclitaxel protein-bound particles); retinoic acid; esperamicins;capecitabine; and pharmaceutically acceptable forms (e.g.,pharmaceutically acceptable salts, hydrates, solvates, isomers,prodrugs, and isotopically labeled derivatives) of any of the above.Also included as suitable chemotherapeutic cell conditioners areanti-hormonal agents that act to regulate or inhibit hormone action ontumors such as anti-estrogens including for example tamoxifen(Nolvadex™), raloxifene, aromatase inhibiting 4(5)-imidazoles,4-hydroxytamoxifen, trioxifene, keoxifene, LY 117018, onapristone, andtoremifene (Fareston); and anti-androgens such as flutamide, nilutamide,bicalutamide, leuprolide, and goserelin; chlorambucil; gemcitabine;6-thioguanine; mercaptopurine; methotrexate; platinum analogs such ascisplatin and carboplatin; vinblastine; platinum; etoposide (VP-16);ifosfamide; mitomycin C; mitoxantrone; vincristine; vinorelbine;navelbine; novantrone; teniposide; daunomycin; aminopterin; xeloda;ibandronate; camptothecin-11 (CPT-11); topoisomerase inhibitor RFS 2000;difluoromethylornithine (DMFO). Where desired, the compounds orpharmaceutical composition as provided herein can be used in combinationwith commonly prescribed anti-cancer drugs such as Herceptin®, Avastin®,Erbitux®, Rituxan®, Taxol®, Arimidex®, Taxotere®, ABVD, AVICINE,abagovomab, acridine carboxamide, adecatumumab,17-N-allylamino-17-demethoxygeldanamycin, alpharadin, alvocidib,3-aminopyridine-2-carboxaldehyde thiosemicarbazone, amonafide,anthracenedione, anti-CD22 immunotoxins, antineoplastic, antitumorigenicherbs, apaziquone, atiprimod, azathioprine, belotecan, bendamustine,BIBW 2992, biricodar, brostallicin, bryostatin, buthionine sulfoximine,CBV (chemotherapy), calyculin, crizotinib, cell-cycle nonspecificantineoplastic agents, dichloroacetic acid, discodermolide,elsamitrucin, enocitabine, epothilone, eribulin, everolimus, exatecan,exisulind, ferruginol, forodesine, fosfestrol, ICE chemotherapy regimen,IT-101, imexon, imiquimod, indolocarbazole, irofulven, laniquidar,larotaxel, lenalidomide, lucanthone, lurtotecan, mafosfamide,mitozolomide, nafoxidine, nedaplatin, olaparib, ortataxel, PAC-1,pawpaw, pixantrone, proteasome inhibitor, rebeccamycin, resiquimod,rubitecan, SN-38, salinosporamide A, sapacitabine, Stanford V,swainsonine, talaporfin, tariquidar, tegafur-uracil, temodar, tesetaxel,triplatin tetranitrate, tris(2-chloroethyl)amine, troxacitabine,uramustine, vadimezan, vinflunine, ZD6126, and zosuquidar.

In some embodiments, the chemotherapeutic is selected from hedgehoginhibitors including, but not limited to IPI-926 (See U.S. Pat. No.7,812,164). Other suitable hedgehog inhibitors include, for example,those described and disclosed in U.S. Pat. No. 7,230,004, U.S. PatentApplication Publication No. 2008/0293754, U.S. Patent ApplicationPublication No. 2008/0287420, and U.S. Patent Application PublicationNo. 2008/0293755, the entire disclosures of which are incorporated byreference herein. Examples of other suitable hedgehog inhibitors includethose described in U.S. Patent Application Publication Nos. US2002/0006931, US 2007/0021493 and US 2007/0060546, and InternationalApplication Publication Nos. WO 2001/19800, WO 2001/26644, WO2001/27135, WO 2001/49279, WO 2001/74344, WO 2003/011219, WO2003/088970, WO 2004/020599, WO 2005/013800, WO 2005/033288, WO2005/032343, WO 2005/042700, WO 2006/028958, WO 2006/050351, WO2006/078283, WO 2007/054623, WO 2007/059157, WO 2007/120827, WO2007/131201, WO 2008/070357, WO 2008/110611, WO 2008/112913, and WO2008/131354, each incorporated herein by reference. Additional examplesof hedgehog inhibitors include, but are not limited to, GDC-0449 (alsoknown as RG3616 or vismodegib) described in, e.g., Von Hoff D. et al.,N. Engl. J. Med. 2009; 361(12):1164-72; Robarge K. D. et al., Bioorg MedChem Lett. 2009; 19(19):5576-81; Yauch, R. L. et al. (2009) Science 326:572-574; Sciencexpress: 1-3 (10.1126/science.1179386); Rudin, C. et al.(2009) New England J of Medicine 361-366 (10.1056/nejma0902903);BMS-833923 (also known as XL139) described in, e.g., in Siu L. et al.,J. Clin. Oncol. 2010; 28:15s (suppl; abstr 2501); and National Instituteof Health Clinical Trial Identifier No. NCT006701891; LDE-225 described,e.g., in Pan S. et al., ACS Med. Chem. Lett., 2010; 1(3): 130-134;LEQ-506 described, e.g., in National Institute of Health Clinical TrialIdentifier No. NCT01106508; PF-04449913 described, e.g., in NationalInstitute of Health Clinical Trial Identifier No. NCT00953758; Hedgehogpathway antagonists disclosed in U.S. Patent Application Publication No.2010/0286114; SMOi2-17 described, e.g., U.S. Patent ApplicationPublication No. 2010/0093625; SANT-1 and SANT-2 described, e.g., inRominger C. M. et al., J. Pharmacol. Exp. Ther. 2009; 329(3):995-1005;1-piperazinyl-4-arylphthalazines or analogues thereof, described inLucas B. S. et al., Bioorg. Med. Chem. Lett. 2010; 20(12):3618-22.

Other hormonal therapy and chemotherapeutic agents include, but are notlimited to, anti-estrogens (e.g. tamoxifen, raloxifene, and megestrolacetate), LHRH agonists (e.g. goserelin and leuprolide), anti-androgens(e.g. flutamide and bicalutamide), photodynamic therapies (e.g.vertoporfin (BPD-MA), phthalocyanine, photosensitizer Pc4, anddemethoxy-hypocrellin A (2BA-2-DMHA)), nitrogen mustards (e.g.cyclophosphamide, ifosfamide, trofosfamide, chlorambucil, estramustine,and melphalan), nitrosoureas (e.g. carmustine (BCNU) and lomustine(CCNU)), alkylsulphonates (e.g. busulfan and treosulfan), triazenes(e.g. dacarbazine, temozolomide), platinum containing compounds (e.g.cisplatin, carboplatin, oxaliplatin), vinca alkaloids (e.g. vincristine,vinblastine, vindesine, and vinorelbine), taxoids or taxanes (e.g.paclitaxel or a paclitaxel equivalent such as nanoparticle albumin-boundpaclitaxel (Abraxane), docosahexaenoic acid bound-paclitaxel(DHA-paclitaxel, Taxoprexin), polyglutamate bound-paclitaxel(PG-paclitaxel, paclitaxel poliglumex, CT-2103, XYOTAX), thetumor-activated prodrug (TAP) ANG1005 (Angiopep-2 bound to threemolecules of paclitaxel), paclitaxel-EC-1 (paclitaxel bound to theerbB2-recognizing peptide EC-1), and glucose-conjugated paclitaxel,e.g., 2′-paclitaxel methyl 2-glucopyranosyl succinate; docetaxel,taxol), epipodophyllins (e.g. etoposide, etoposide phosphate,teniposide, topotecan, 9-aminocamptothecin, camptoirinotecan,irinotecan, crisnatol, mytomycin C), anti-metabolites, DHFR inhibitors(e.g. methotrexate, dichloromethotrexate, trimetrexate, edatrexate), IMPdehydrogenase inhibitors (e.g. mycophenolic acid, tiazofurin, ribavirin,and EICAR), ribonuclotide reductase inhibitors (e.g. hydroxyurea anddeferoxamine), uracil analogs (e.g. 5-fluorouracil (5-FU), floxuridine,doxifluridine, raltitrexed, tegafur-uracil, capecitabine), cytosineanalogs (e.g. cytarabine (ara C, cytosine arabinoside), andfludarabine), purine analogs (e.g. mercaptopurine and thioguanine),Vitamin D3 analogs (e.g. EB 1089, CB 1093, and KH 1060), isoprenylationinhibitors (e.g. lovastatin), dopaminergic neurotoxins (e.g.1-methyl-4-phenylpyridinium ion), cell cycle inhibitors (e.g.staurosporine), actinomycin (e.g. actinomycin D, dactinomycin),bleomycin (e.g. bleomycin A2, bleomycin B2, peplomycin), anthracyclines(e.g. daunorubicin, doxorubicin, pegylated liposomal doxorubicin,idarubicin, epirubicin, pirarubicin, zorubicin, mitoxantrone), MDRinhibitors (e.g. verapamil), Ca2+|ATPase inhibitors (e.g. thapsigargin),thalidomide, lenalidomide (REVLIMID®), tyrosine kinase inhibitors (e.g.,axitinib (AG013736), bosutinib (SKI-606), cediranib (RECENTIN™,AZD2171), dasatinib (SPRYCEL®, BMS-354825), erlotinib (TARCEVA®),gefitinib (IRESSA®), imatinib (Gleevec®, CGP57148B, STI-571), lapatinib(TYKERB®, TYVERB®), lestaurtinib (CEP-701), neratinib (HKI-272),nilotinib (TASIGNA®), semaxanib (semaxinib, SU5416), sunitinib (SUTENT®,SU11248), toceranib (PALLADIA®), vandetanib (ZACTIMA®, ZD6474),vatalanib (PTK787, PTK/ZK), trastuzumab (HERCEPTIN®), bevacizumab(AVASTIN®), rituximab (RITUXAN®), cetuximab (ERBITUX®), panitumumab(VECTIBIX®), ranibizumab (Lucentis®), sorafenib (NEXAVAR®), everolimus(AFINITOR®), alemtuzumab (CAMPATH®), gemtuzumab ozogamicin (MYLOTARG®),temsirolimus (TORISEL®), ENMD-2076, PCI-32765, AC220, dovitinib lactate(TKI258, CHIR-258), BIBW 2992 (TOVOK™), SGX523, PF-04217903,PF-02341066, PF-299804, BMS-777607, ABT-869, MP470, BIBF 1120(VARGATEF®), AP24534, JNJ-26483327, MGCD265, DCC-2036, BMS-690154,CEP-11981, tivozanib (AV-951), OSI-930, MM-121, XL-184, XL-647, and/orXL228), proteasome inhibitors (e.g., bortezomib (Velcade)), mTORinhibitors (e.g., rapamycin, temsirolimus (CCI-779), everolimus(RAD-001), ridaforolimus, AP23573 (Ariad), AZD8055 (AstraZeneca), BEZ235(Novartis), BGT226 (Norvartis), XL765 (Sanofi Aventis), PF-4691502(Pfizer), GDC0980 (Genetech), SF1126 (Semafoe) and OSI-027 (OSI)),oblimersen, gemcitabine, carminomycin, leucovorin, pemetrexed,cyclophosphamide, dacarbazine, procarbazine, prednisolone,dexamethasone, camptothecin, plicamycin, asparaginase, aminopterin,methopterin, porfiromycin, melphalan, leurosidine, leurosine,chlorambucil, trabectedin, procarbazine, discodermolide, carminomycin,aminopterin, and hexamethyl melamine.

Exemplary biotherapeutic agents include, but are not limited to,interferons, cytokines (e.g., tumor necrosis factor, interferon α,interferon γ), vaccines, hematopoietic growth factors, monoclonalserotherapy, immuno-stimulants and/or immuno-modulatory agents (e.g.,IL-1, 2, 4, 6, 7, 12, 15, or 21), immune cell growth factors (e.g.,GM-CSF) and antibodies (e.g. Herceptin (trastuzumab), T-DM1, AVASTIN(bevacizumab), ERBITUX (cetuximab), Vectibix (panitumumab), Rituxan(rituximab), Bexxar (tositumomab), or Perjeta (pertuzumab)).

In some embodiments, the biotherapeutic agent is an immunotherapeuticagent, e.g., a cancer vaccine e.g., a tumor vaccine. Exemplary cancervaccines include Aduro (GVAX); Advaxis (ADXS11-001, ADXS31-001,ADXS31-164, ADXS31-142 (ADXS-PSA)); ALVAC-CEA vaccine; Avax Technologies(AC Vaccine); Amgen (talimogene laherparepvec); Biovest International(BiovaxID in phase III); Bavarian Nordic (PROSTVAC); CelldexTherapeutics (CDX110, CDX1307 and CDX1401); The Center of MolecularImmunology (CimaVax-EGF); CureVac develops mRNA-based cancerimmunotherapies; CV9104; Dendreon Corp (Neuvenge); Galena Biopharma(NeuVax); Antigen Express (Ae-37); Geron Corporation (GRNVAC1);GlobeImmune (Tarmogens, GI-4000, GI-6207, GI-6301); Heat Biologics(ImPACT Therapy); Immatics biotechnologies (IMA901); Merck (Stimuvax);Panacela Labs, Inc. (MOBILAN Adenovirus-based treatment); Prima BioMed(Cvac); Scancell Holdings (SCIB1).

In embodiments, the biotherapeutic agent is a cellular therapy, e.g.,dendritic cell therapy or a chimeric T cell therapy such as CART.Dendritic cell therapy can comprise loading dendritic cells with anantigen obtained from a patient's tumor, then administering thedendritic cells to the patient in order to sensitize the patient's own Tcells to the tumor antigens. Chimeric antigen receptors (CARs) areengineered receptors that can be used to confer tumor specificity to a Tcell. CARS have been generated with specificity for α-folate receptor,CAIX, CD19, CD20, CD22, CD30, CD33, CD44v7/8, CEA, EGP-2, EGP-40,erb-B2, erb-B 2,3,4, FBP, Fetal acethylcholine receptor, FD2, Her2/neu,IL13R-a2, KDR, k-light chain, LeY, L1 cell adhesion molecule, MAGE-A1,mesothelin, CMV-infected cells, MUC1, NKG2D ligands, oncofetal antigenh5T4, PSCA, PSMA, TAA, TAG-72, and VEGF-R2.

In one embodiment, the biotherapeutic agent is an anti-CD37 antibodysuch as, but not limited to, IMGN529, K7153A and TRU-016. In anotherembodiment, the biotherapeutic agent is an anti-CD20 antibody such as,but not limited to, ¹³¹I tositumomab, ⁹⁰Y ibritumomab, ¹¹¹I ibritumomab,obinutuzumab (GAZYVA), and ofatumumab. In another embodiment, thebiotherapeutic agent is an anti-CD52 antibody such as, but not limitedto, alemtuzumab.

In some embodiments, the chemotherapeutic is selected from HSP90inhibitors. The HSP90 inhibitor can be a geldanamycin derivative, e.g.,a benzoquinone or hygroquinone ansamycin HSP90 inhibitor (e.g., IPI-493and/or IPI-504). Non-limiting examples of HSP90 inhibitors includeIPI-493, IPI-504, 17-AAG (also known as tanespimycin or CNF-1010),BIIB-021 (CNF-2024), BIIB-028, AUY-922 (also known as VER-49009),SNX-5422, STA-9090, AT-13387, XL-888, MPC-3100, CU-0305, 17-DMAG,CNF-1010, Macbecin (e.g., Macbecin I, Macbecin II), CCT-018159,CCT-129397, PU-H71, or PF-04928473 (SNX-2112).

In some embodiments, the chemotherapeutic is selected from PI3Kinhibitors (e.g., including those PI3K inhibitors provided herein andthose PI3K inhibitors not provided herein). In some embodiment, the PI3Kinhibitor is an inhibitor of delta and gamma isoforms of PI3K. In someembodiment, the PI3K inhibitor is an inhibitor of delta isoform of PI3K.In some embodiment, the PI3K inhibitor is an inhibitor of gamma isoformof PI3K. In some embodiments, the PI3K inhibitor is an inhibitor ofalpha isoform of PI3K. In other embodiments, the PI3K inhibitor is aninhibitor of one or more alpha, beta, delta and gamma isoforms of PI3K.Exemplary PI3K inhibitors that can be used in combination are describedin, e.g., WO 09/088990, WO 09/088086, WO 2011/008302, WO 2010/036380, WO2010/006086, WO 09/114870, WO 05/113556; US 2009/0312310, and US2011/0046165, each incorporated herein by reference. Additional PI3Kinhibitors that can be used in combination with the pharmaceuticalcompositions, include but are not limited to, RP-6530, TG 100-115,RV1729, AMG-319, GSK 2126458, GDC-0980, GDC-0941, Sanofi XL147, XL499,XL756, XL147, PF-4691502, BKM 120, CAL-101 (GS-1101), CAL 263, SF1126,PX-886, and a dual PI3K inhibitor (e.g., Novartis BEZ235). In oneembodiment, the PI3K inhibitor is an isoquinolinone. In one embodiment,the PI3K inhibitor is RP-6530, which has the chemical name:(S)-2-(1-((9H-purin-6-yl)amino)propyl)-3-(3-fluorophenyl)-4H-chromen-4-one.In one embodiment, the PI3K inhibitor is TG 100-115, which has thechemical name: 6,7-Bis(3-hydroxyphenyl)pteridine-2,4-diamine. In oneembodiment, the PI3K inhibitor is RV1729, which has the chemical name:6-(2-((4-amino-3-(3-hydroxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl)-3-(2-chlorobenzyl)-4-oxo-3,4-dihydroquinazolin-5-yl)-N,N-bis(2-methoxyethyl)hex-5-ynamide.

Without being bound by any particular theory, it is believed that therole of each PI3K isoform is critically dependent on cell type andupstream initiating signals, and therefore pharmacologic inhibition ofspecific isoforms can lead to different physiologic outcomes. PI3K is alipid kinase existing in multiple isoforms that have central roles inthe regulation of important cellular processes, including cell growthand survival. Puri et al., Frontiers in Immunology. 2012, 3: 256. PI3K-δand PI3K-γ are both expressed in CLL and NHL tumor cells. Signalingthrough PI3K is critical for supporting the growth and survival of thesemalignancies as they mediate intracellular BCR signaling and promoteinteractions between the tumor cells and their microenvironment. Puri etal., Frontiers in Immunology. 2012, 3: 256.

The specific functions of PI3K-δ in malignant B cells support therationale for it to be a therapeutic target to control these diseases.PI3K-δ inhibition disrupts malignant cell interaction with the stromalmicroenvironment, thereby short-circuiting chemokine-mediatedstimulation of CLL and other B-cell malignancies, priming cells forapoptosis by pharmacologic or natural stimuli. Pharmacologic inhibitionof PI3K-δ reduces disease activity in various models of B-cell-derivedmalignancies, including CLL and B-cell lymphomas. PI3K-δ inhibitionimproves the therapeutic potential of other antitumor agents in variouspreclinical models of B-cell malignancy, including CLL. Lannutti et al.,Blood. 2011, 117, 591-594.

The role of PI3K-γ in cells that maintain the malignant B-cellmicroenvironment creates potential for therapeutic inhibition of PI3K-γto control these diseases. PI3K-γ plays a role in T-cell activation andmigration and GPCR-associated chemokine signaling. Reif et al. JImmunol. 2004; 173:2236-2240. PI3K-γ also mediates adhesion andtrafficking of tumor-associated macrophages (TAMs). Reif et al. JImmunol. 2004; 173:2236-2240. Hasan et al., Int Immunopharmacology.2010, 10, 1017-1021; Laffargue et al., Immunity. 2002, 16, 441-451.There is dynamic interplay or “cross-talk” between PI3K-δ and PI3K-γ inessential cellular activities in malignant cells. In certain tumortypes, PI3K-γ can promote tumorigenesis in the absence of PI3K-δ.Subramaniam et al., Cancer Cell. 2012, 21, 459-472.

In certain tumor types, dual isoform inhibition may be necessary foroptimal tumor growth inhibition in preclinical models. Subramaniam etal., Cancer Cell. 2012, 21, 459-472. As shown in the examples, in somecell lines (e.g., NHL (e.g., follicular lymphoma), DLBCL, mantle cell,multiple myeloma, T-cell lymphoma), combined inhibition of PI3K-δ andPI3K-γ shows greater growth inhibition than inhibition of either isoformalone.

In certain embodiments, provided herein are pharmaceutical compositionscomprising a PI3K gamma selective compound and a PI3K delta compound ora PI3K delta selective compound. In one embodiment, the composition issynergistic in treating or preventing a PI3K mediated disorder.

Also provided herein are methods of treating or preventing a PI3Kmediated disorder in a subject, comprising administering to the subjecta therapeutically effective amount of a PI3K gamma selective compound incombination with a PI3K delta compound or a PI3K delta selectivecompound. Also provided herein are methods of enhancing a PI3K deltacompound treatment of a PI3K mediated disorder in a subject comprisingadministering a PI3K gamma selective compound in combination with thePI3K delta compound. In one embodiment, the administering the PI3K gammaselective compound in combination with the PI3K delta compound providessynergistic effect. In one embodiment, the administering the PI3K gammaselective compound in combination with the PI3K delta compound providesadditive effect. In another embodiment, administering the PI3K gammaselective compound in combination with the PI3K delta compound providesa faster response time compared to administering a delta selectivecompound alone.

Also provided herein are methods for inhibiting migration of peripheralT-cells in a subject diagnosed with CLL comprising administering a PI3Kgamma selective compound to the subject.

Also provided herein are methods of inhibiting growth of a cellcomprising contacting the cell with a PI3K gamma selective compound incombination with a PI3K delta selective compound. In one embodiment, thecell is a cancer cell. In another embodiment, the cell is in a subject.In one embodiment, the subject is afflicted with a proliferativedisease, cancer, autoimmune disease, or inflammatory disease.

In one embodiment, the PI3K gamma selective compound selectivelyinhibits PI3K gamma isoform over PI3K delta isoform. In one embodiment,the PI3K gamma selective compound has a delta/gamma selectivity ratio ofgreater than 1, greater than about 5, greater than about 10, greaterthan about 50, greater than about 100, greater than about 200, greaterthan about 400, greater than about 600, greater than about 800, greaterthan about 1000, greater than about 1500, greater than about 2000,greater than about 5000, greater than about 10,000, or greater thanabout 20,000. In one embodiment, the PI3K gamma selective compound has adelta/gamma selectivity ratio in the range of from greater than 1 toabout 5, from about 5 to about 10, from about 10 to about 50, from about50 to about 850, or greater than about 850. In one embodiment, thedelta/gamma selectivity ratio is determined by dividing the compound'sIC₅₀ against PI3K delta isoform by the compound's IC₅₀ against PI3Kgamma isoform.

In one embodiment, the PI3K gamma selective compound is a compoundprovided herein, e.g., a compound of Formula (I′), (A′), (I), or (A), ora pharmaceutically acceptable form thereof. In one embodiment, the PI3Kgamma selective compound is a compound of Formula (I′), (A′), (I), or(A), or a pharmaceutically acceptable form thereof, wherein R¹ is aheteroaryl (e.g., a 5-membered heteroaryl or a 6-membered heteroaryl),In one embodiment, the PI3K gamma selective compound is Compound 2, 4,7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63,73, 75, 77, 79, 80, 81, or 88, or a pharmaceutically acceptable formthereof. In one embodiment, the PI3K gamma selective compound isCompound 4, 7, 9, 19, 27, 35, 37, 41, 52, 73, 80, or 88, or apharmaceutically acceptable form thereof.

In one embodiment, the PI3K delta selective compound selectivelyinhibits PI3K delta isoform over PI3K gamma isoform. In one embodiment,the PI3K delta selective compound has a gamma/delta selectivity ratio ofgreater than 1, greater than about 5, greater than about 10, greaterthan about 50, greater than about 100, greater than about 200, greaterthan about 400, greater than about 600, greater than about 800, greaterthan about 1000, greater than about 1500, greater than about 2000,greater than about 5000, greater than about 10,000, or greater thanabout 20,000. In one embodiment, the PI3K delta selective compound has agamma/delta selectivity ratio in the range of from greater than 1 toabout 5, from about 5 to about 10, from about 10 to about 50, from about50 to about 850, or greater than about 850. In one embodiment, thegamma/delta selectivity ratio is determined by dividing the inhibitor'sIC₅₀ against PI3K gamma isoform by the inhibitor's IC₅₀ against PI3Kdelta isoform.

In certain embodiments, the PI3K inhibitor is a PI3K delta selectivecompound. In one embodiment, the PI3K delta selective compoundselectively inhibits PI3K delta isoform over PI3K alpha isoform. In oneembodiment, the PI3K delta selective compound has an alpha/deltaselectivity ratio of greater than 1, greater than about 5, greater thanabout 10, greater than about 50, greater than about 100, greater thanabout 200, greater than about 400, greater than about 600, greater thanabout 800, greater than about 1000, greater than about 1500, greaterthan about 2000, greater than about 5000, greater than about 10,000, orgreater than about 20,000. In one embodiment, the PI3K delta selectivecompound has an alpha/delta selectivity ratio in the range of fromgreater than 1 to about 5, from about 5 to about 10, from about 10 toabout 50, from about 50 to about 850, or greater than about 850. In oneembodiment, the alpha/delta selectivity ratio is determined by dividingthe inhibitor's IC₅₀ against PI3K alpha isoform by the inhibitor's IC₅₀against PI3K delta isoform.

In certain embodiments, the PI3K inhibitor is a PI3K delta selectivecompound. In one embodiment, the PI3K delta selective compoundselectively inhibits PI3K delta isoform over PI3K beta isoform. In oneembodiment, the PI3K delta selective compound has a beta/deltaselectivity ratio of greater than 1, greater than about 5, greater thanabout 10, greater than about 50, greater than about 100, greater thanabout 200, greater than about 400, greater than about 600, greater thanabout 800, greater than about 1000, greater than about 1500, greaterthan about 2000, greater than about 5000, greater than about 10,000, orgreater than about 20,000. In one embodiment, the PI3K delta selectivecompound has a beta/delta selectivity ratio in the range of from greaterthan 1 to about 5, from about 5 to about 10, from about 10 to about 50,from about 50 to about 850, or greater than about 850. In oneembodiment, the beta/delta selectivity ratio is determined by dividingthe compound's IC₅₀ against PI3K beta isoform by the compound's IC₅₀against PI3K delta isoform.

In one embodiment, the PI3K delta selective compound is GSK-2269557(2-(6-(1H-indol-4-yl)-1H-indazol-4-yl)-5-((4-isopropylpiperazin-1-yl)methyl)oxazole),GS9820 (CAL-120,(S)-2-(1-((9H-purin-6-yl)amino)ethyl)-6-fluoro-3-phenylquinazolin-4(3H)-one),GS-1101(5-fluoro-3-phenyl-2-([S)]-1-[9H-purin-6-ylamino]-propyl)-3H-quinazolin-4-one),AMG319, or TGR-1202((S)-2-(1-(4-amino-3-(3-fluoro-4-isopropoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-6-fluoro-3-(3-fluorophenyl)-4H-chromen-4-one),or a mixture thereof. In one embodiment, the PI3K delta selectivecompound is GS1101.

In one embodiment, the PI3K delta selective compound is a PI3K deltainhibitor as described in WO 2005/113556, the entirety of which isincorporated herein by reference. In one embodiment, the PI3K deltaselective compound has an alpha/delta IC₅₀ ratio of greater than about200, greater than about 250, greater than about 300, greater than about500, greater than about 600, or greater than about 700. In oneembodiment, the PI3K delta selective compound has a beta/delta IC₅₀ratio of greater than about 50, greater than about 75, greater thanabout 80, greater than about 90, or greater than about 100. In oneembodiment, the PI3K delta selective compound has a gamma/delta IC₅₀ratio of greater than about 50, greater than about 75, greater thanabout 80, greater than about 100, greater than about 200, greater thanabout 300, greater than about 400, or greater than about 500. In oneembodiment, the PI3K delta selective compound is Compound No. 113 or 107as described in 2005/113556.

In one embodiment, the PI3K delta selective compound is a PI3K deltacompound as described in WO2014/006572, the entirety of which isincorporated herein by reference. In one embodiment, the PI3K deltaselective compound is a PI3K delta inhibitor has an alpha/deltaselectivity ratio of greater than about 100, greater than about 250,greater than about 500, greater than about 750, or greater than about1000. In one embodiment, the PI3K delta selective compound is a PI3Kdelta inhibitor has a beta/delta selectivity ratio of greater than about10, greater than about 20, greater than about 30, greater than about 40,or greater than about 50. In one embodiment, the PI3K delta selectivecompound is a PI3K delta inhibitor has a gamma/delta selectivity ratioof greater than about 1, greater than about 10, greater than about 25,greater than about 30, or greater than about 50. In one embodiment, thePI3K delta selective compound is Compound Nos. A1, A2, B, B1 or B2 asdescribed in WO2014/006572. In one embodiment, the PI3K delta selectivecompound is Compound No. B1 as described in WO2014/006572.

In one embodiment, the PI3K delta selective compound is a PI3K deltacompound as described in WO 2013/032591, the entirety of which isincorporated herein by reference. In one embodiment, the PI3K deltaselective compound is a compound of Formula (I) as described in WO2013/032591. In one embodiment, the PI3K delta selective compound is acompound described in WO 2013/032591 with a IC₅₀ (nM) for the PI3K deltaisoform of less than 100 nM and a IC₅₀ (nM) for the PI3K alpha, beta, orgamma of greater than about 100 nM, greater than about 1 μM, or greaterthan about 10 μM. In one embodiment, the PI3K delta selective compoundhas an alpha/delta selectivity ratio, a beta/delta selectivity ratio, ora gamma/delta selectivity ratio of greater than 1, greater than about10, or greater than about 100. In one embodiment, the PI3K deltaselective compound is Compound Nos. 13, 30, 41, 55, 57, 124, 167, 183,185, 187, 191, 196, 226, 230, 232, 234, 235, 326, 327, 328, 333, 334,336, 337, 338, 356, 359, 378, 439, 440, 443, or 455, as described in WO2013/032591. In one embodiment, the PI3K delta selective compound isCompound Nos. 183, 230, 234, 235, 326, 333, 336, 337, 338, or 359, asdescribed in WO 2013/032591. In one embodiment, the PI3K delta selectivecompound is Compound No. 359 as described in WO 2013/032591.

In one embodiment, provided herein are pharmaceutical compositionscomprising a compound of Formula (I′), (A′), (I), or (A), or apharmaceutically acceptable form thereof, and a PI3K delta selectivecompound, wherein the PI3K delta selective compound is GSK-2269557,GS-9820, GS-1101 (Cal-101 or idelalisib), AMG319, or TGR-1202, or amixture thereof. In one embodiment, the PI3K delta selective compound isGS1101. In one embodiment, the composition is synergistic in treating orpreventing a PI3K mediated disorder. In one embodiment, the PI3K deltaselective compound is a compound described in WO2011/146882, theentirety of which is incorporated herein by reference. In oneembodiment, the PI3K delta selective compound is a compound described inWO2011/146882 with a IC₅₀ (nM) for the PI3K delta isoform of less than100 nM and a IC₅₀ (nM) for the PI3K alpha, beta, or gamma of greaterthan about 100 nM, greater than about 1 μM, or greater than about 10 μM.In one embodiment, the PI3K delta selective compound has an alpha/deltaselectivity ratio, a beta/delta selectivity ratio, or a gamma/deltaselectivity ratio of greater than 1, greater than about 10, or greaterthan about 100. In one embodiment, the PI3K delta selective compound isCompound No. 69 as described in WO2011/146882.

In one embodiment, the PI3K delta selective compound is a compounddescribed in WO2013/012915, the entirety of which is incorporated hereinby reference. In one embodiment, the PI3K delta selective compound is acompound described in WO2013/012915 with a IC₅₀ (nM) for the PI3K deltaisoform of less than 100 nM and a IC₅₀ (nM) for the PI3K alpha, beta, orgamma of greater than about 100 nM, greater than about 1 μM, or greaterthan about 10 μM. In one embodiment, the PI3K delta selective compoundhas an alpha/delta selectivity ratio, a beta/delta selectivity ratio, ora gamma/delta selectivity ratio of greater than 1, greater than about10, or greater than about 100. In one embodiment, the PI3K deltaselective compound is Compound No. 1-41 or 1-106 as described inWO2013/012915.

In one embodiment, the PI3K delta selective compound is a compounddescribed in WO2013/012918, the entirety of which is incorporated hereinby reference. In one embodiment, the PI3K delta selective compound is acompound described in WO2013/012918 with a IC₅₀ (nM) for the PI3K deltaisoform of less than 100 nM and a IC₅₀ (nM) for the PI3K alpha, beta, orgamma of greater than about 100 nM, greater than about 1 μM, or greaterthan about 10 μM. In one embodiment, the PI3K delta selective compoundhas an alpha/delta selectivity ratio, a beta/delta selectivity ratio, ora gamma/delta selectivity ratio of greater than 1, greater than about10, or greater than about 100. In one embodiment, the PI3K deltaselective compound is Compound No. 19, 28, 37, 38, 51, 59, 60, 89, 92,103, 106, 107, 108, or 109 as described in WO2013/012918. In oneembodiment, the PI3K delta selective compound is Compound No. 103 or 106as described in WO2013/012918.

In one embodiment, provided herein are methods of treating or preventinga PI3K mediated disorder in a subject, comprising administering to thesubject a therapeutically effective amount of a compound of Formula(I′), (A′), (I), or (A), or a pharmaceutically acceptable form thereof,in combination with a PI3K delta selective compound, wherein the PI3Kdelta selective compound is GSK-2269557, GS-9820, GS-1101 (Cal-101),AMG319, or TGR-1202, or a mixture thereof. In one embodiment, providedherein are methods of enhancing a PI3K delta selective compoundtreatment of a PI3K mediated disorder in a subject comprisingadministering a compound of Formula (I′), (A′), (I), or (A), or apharmaceutically acceptable form thereof, in combination with the PI3Kselective delta compound, wherein the PI3K delta selective compound isGSK-2269557, GS9-820, GS-1101 (Cal-101), AMG319, or TGR-1202, or amixture thereof. In one embodiment, the PI3K delta selective compound isGS1101. In one embodiment, the administering a compound of Formula (I′),(A′), (I), or (A), or a pharmaceutically acceptable form thereof, incombination with the PI3K delta selective compound provides synergisticeffect.

Also provided herein are methods of inhibiting growth of a cellcomprising contacting the cell with a compound of Formula (I′), (A′),(I), or (A), or a pharmaceutically acceptable form thereof, incombination with a PI3K delta selective compound, wherein the PI3K deltaselective compound is GSK-2269557, GS-9820, GS-1101 (Cal-101), AMG319,or TGR-1202, or a mixture thereof. In one embodiment, the PI3K deltaselective compound is GS1101. In one embodiment, the cell is a cancercell. In another embodiment, the cell is in a subject. In oneembodiment, the subject is afflicted with a proliferative disease,cancer, autoimmune disease, or inflammatory disease.

In one embodiment, the PI3K delta selective compound is a compoundselected from US Patent Publication Nos. 20140058103, 20140051699,20140045825, 20140011819, 20130231356, 20130225557, 20120245144,20100305084, 20100256167, 20100168139, 20100152211, and 20100029693. Inone embodiment, the PI3K delta selective compound is a compound selectedfrom U.S. Pat. Nos. 8,653,077, 8,637,533, 8,623,881, 8,586,597,8,569,296, 8,563,540, 8,492,389, 8,440,651, 8,138,195, 7,932,260, and6,949,535.

For example, a compound provided herein with a delta/gamma selectivityratio of greater than 150 can be combined with a compound that has agamma/delta selectivity ratio of 1000 at various amounts (e.g., a ratioof 10:1 or 40:1 of a gamma selective compound and a delta selectivecompound) to provide synergistic effect in cell lines (e.g., diffuselarge B-cell lymphoma cell lines such as SU-DHL-4,TMD-8 and Farage).

The PI3K gamma selective compound and PI3K delta selective compoundcomposition or combination therapy can provide synergistic effect intreating or preventing a PI3K mediated disorder. In one embodiment, thedisorder is a cancer. In one embodiment, the cancer is diffuse largeB-cell lymphoma (e.g., TMD-8 and Farage cell lines), B-cell lymphoma(e.g., karpas-422 cell line), T-cell lymphoma, non-Hodgkin's lymphoma,Hodgkin lymphoma, or anaplastic large cell lymphoma (e.g., HH cellline).

In some embodiments, the synergistic effect can be characterized by anisobologram. Potency shifting is usually shown using an isobologramwhich shows how much less a compound is required in combination toachieve a desired effect level, when compared to the single agent dosesneeded to reach that effect. The choice of effect level for theisobologram display and combination index calculations can either bemanually or automatically selected in the Chalice Analyzer. Potencyshifting is scored as the combination index (CI). Chou et al., AdvEnzyme Regal 1984; 22: 27-55. The CI is a rough estimate of how much acompound was needed in combination relative to the single agent dosesrequired to achieve the chosen effect level, and a value of 0.1 meansthat only a tenth of equivalent amounts of the single agents were neededfor the combination to reach the same effect level. Additive effect isCI=1.0. Synergistic effect is CI<1. Antagonistic effect is CI>1.0.

In some embodiment, the synergisitic effect is characterized by SynergyScore.

Different stimuli can be used to preferentially induce T-cell orCLL-cell migration. For example, CCL19 and CCL21 stimuli selectivelyinduce migration of both CLL and T-cells. CXCL13 is CLL-cell specific,whereas CXCL12 is T-cell specific. As such, stimuli CXCL13 and CXCL12can be used to induce CLL-cell and T-cell migrations, respectively. ThePI3K gamma selective compounds provided herein can inhibitcancer-promoting cell migration, e.g., CXCL 12-induced T-cell migration.In some embodiments, elevated pAKT levels indicate that theCXCL12-induced migration machinery is activated. Consequently, in someembodiments, the PI3K gamma selective compound, e.g., Compound 4,interferes with AKT signaling and/or reduces pAKT levels in the T-cells.In one embodiment, the PI3K gamma selective compound is a compound thathas a delta/gamma selectivity ratio of greater than about 50, such asCompound 4. In another embodiment, the PI3K delta selective compound isa compound that has a gamma/delta selectivity ratio of greater thanabout 50. The gamma selective compound can be more potent than a deltaselective compound at inhibiting cancer-promoting cell migration, e.g.,CXCL12-induced T cell migration in CLL PBMCs. The ability to inhibit themigration of cancer-promoting cells can stop the growth of cancers byblocking the migration of cells that promote cancer growth to the cancercell niche. In another embodiment, gamma or delta selective compoundscan inhibit the migration of cancer cells themselves and limit cancercell dissemination. As such, the gamma selective compounds providedherein can be used to treat and/or prevent cancer, or slow down theprogression of cancer or metastasis. Treatment with a combination ofgamma and delta selective compounds can have an earlier response timecompared to a delta selective compound alone, for example in B cellmeditated cancers.

In one embodiment, a compound provided herein (e.g., a compound of anyof Formulae (I″), (I′), (A′), (I), (A), (II), (III), (IV), (V), (VI),(VII), (IX), (X), (XI), (XII), (XIII), (XIV), (XV), (XVI), and (XVII),e.g., a selective PI3K-γ inhibitor, e.g., Compound 4), or apharmaceutically acceptable form thereof, is administered in combinationwith a chemotherapy (e.g., temozolomide) for the treatment of a cancer.In one embodiment, the cancer is glioblastoma (e.g., glioblastomamultiforme). In one embodiment, provided herein is a method of treatingglioblastoma in a subject, comprising administering to the subject atherapeutically effective amount of Compound 4, or a pharmaceuticallyacceptable form thereof, in combination with a chemotherapy. In oneembodiment, the compound is administered subsequent to the chemotherapy.In one embodiment, the compound is administered concurrently to thechemotherapy. In one embodiment, the compound is administered prior tothe chemotherapy.

In some embodiments, provided herein is a method for using a compoundprovided herein, or a pharmaceutically acceptable form (e.g.,pharmaceutically acceptable salts, hydrates, solvates, isomers,prodrugs, and isotopically labeled derivatives) thereof, or apharmaceutical composition as provided herein, in combination withradiation therapy in inhibiting abnormal cell growth or treating thehyperproliferative disorder in the subject. Techniques for administeringradiation therapy are known in the art, and these techniques can be usedin the combination therapy described herein. The administration of acompound provided herein in this combination therapy can be determinedas described herein.

In certain embodiments, provided herein are methods of treating a solidtumor in a subject, comprising administering to the subject atherapeutically effective amount of a compound provided herein (e.g., acompound of any of Formulae (I″), (I′), (A′), (I), (A), (II), (III),(IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII), (XIV), (XV),(XVI), and (XVII), e.g., a selective PI3K-γ inhibitor, e.g., Compound4), or a pharmaceutically acceptable form thereof, in combination with aradiation therapy.

In one embodiment, the solid tumor is selected from one or more of: acancer of the pulmonary system, a brain cancer, a cancer of thegastrointestinal tract, a skin cancer, a genitourinary cancer, apancreatic cancer, a lung cancer, a medullobastoma, a basal cellcarcinoma, a glioma, a breast cancer (e.g., triple negative breastcancer), a prostate cancer, a testicular cancer, an esophageal cancer, ahepatocellular cancer, a gastric cancer, a gastrointestinal stromaltumor (GIST), a colon cancer, a colorectal cancer, an ovarian cancer, amelanoma, a neuroectodermal tumor, head and neck cancer, a sarcoma, asoft-tissue sarcoma, fibrosarcoma, myxosarcoma, liposarcoma, achondrosarcoma, an osteogenic sarcoma, a chordoma, an angiosarcoma, anendotheliosarcoma, a lymphangiosarcoma, a lymphangioendotheliosarcoma, asynovioma, a mesothelioma, a leiomyosarcoma, a cervical cancer, auterine cancer, an endometrial cancer, a carcinoma, a bladder carcinoma,an epithelial carcinoma, a squamous cell carcinoma, an adenocarcinoma, abronchogenic carcinoma, a renal cell carcinoma, a hepatoma, a bile ductcarcinoma, a neuroendocrine cancer, a carcinoid tumor, diffuse typegiant cell tumor, andglioblastoma.

In one embodiment, the compound, or a pharmaceutically acceptable formthereof, is administered after the radiation therapy is administered. Inone embodiment, the compound, or a pharmaceutically acceptable formthereof, is administered at the same time that radiation therapy isadministered. In one embodiment, the compound, or a pharmaceuticallyacceptable form thereof, is administered alone after discontinuing theradiation therapy.

Radiation therapy can be administered through one of several methods, ora combination of methods, including without limitation, external-beamtherapy, internal radiation therapy, implant radiation, stereotacticradiosurgery, systemic radiation therapy, radiotherapy and permanent ortemporary interstitial brachytherapy. The term “brachytherapy,” as usedherein, refers to radiation therapy delivered by a spatially confinedradioactive material inserted into the body at or near a tumor or otherproliferative tissue disease site. The term is intended withoutlimitation to include exposure to radioactive isotopes (e.g., At-211,1-131, 1-125, Y-90, Re-186, Re-188, Sm-153, Bi-212, P-32, andradioactive isotopes of Lu). Suitable radiation sources for use as acell conditioner as provided herein include both solids and liquids. Byway of non-limiting example, the radiation source can be a radionuclide,such as 1-125, 1-131, Yb-169, Ir-192 as a solid source, 1-125 as a solidsource, or other radionuclides that emit photons, beta particles, gammaradiation, or other therapeutic rays. The radioactive material can alsobe a fluid made from any solution of radionuclide(s), e.g., a solutionof 1-125 or 1-131, or a radioactive fluid can be produced using a slurryof a suitable fluid containing small particles of solid radionuclides,such as Au-198, Y-90. Moreover, the radionuclide(s) can be embodied in agel or radioactive micro spheres.

Without being limited by any theory, a compound provided herein, or apharmaceutically acceptable form (e.g., pharmaceutically acceptablesalts, hydrates, solvates, isomers, prodrugs, and isotopically labeledderivatives) thereof, or a pharmaceutical composition as providedherein, can render abnormal cells more sensitive to treatment withradiation for purposes of killing and/or inhibiting the growth of suchcells. Accordingly, provided herein is a method for sensitizing abnormalcells in a subject to treatment with radiation which comprisesadministering to the subject an amount of a compound provided herein, ora pharmaceutically acceptable form (e.g., pharmaceutically acceptablesalts, hydrates, solvates, isomers, prodrugs, and isotopically labeledderivatives) thereof, which amount is effective in sensitizing abnormalcells to treatment with radiation. The amount of the compound used inthis method can be determined according to the means for ascertainingeffective amounts of such compounds described herein.

In one embodiment, a compound as provided herein, or a pharmaceuticallyacceptable form (e.g., pharmaceutically acceptable salts, hydrates,solvates, isomers, prodrugs, and isotopically labeled derivatives)thereof, or a pharmaceutical composition as provided herein, can be usedin combination with an amount of one or more substances selected fromanti-angiogenesis agents, signal transduction inhibitors, andantiproliferative agents, glycolysis inhibitors, or autophagyinhibitors.

Other therapeutic agents, such as MMP-2 (matrix-metalloproteinase 2)inhibitors, MMP-9 (matrix-metalloproteinase 9) inhibitors, and COX-11(cyclooxygenase 11) inhibitors, can be used in conjunction with acompound provided herein, or a pharmaceutically acceptable form thereof,or a pharmaceutical composition described herein. Such therapeuticagents include, for example, rapamycin, temsirolimus (CCI-779),everolimus (RAD001), sorafenib, sunitinib, and bevacizumab. Examples ofuseful COX-II inhibitors include CELEBREX™ (alecoxib), valdecoxib, androfecoxib. Examples of useful matrix metalloproteinase inhibitors aredescribed in WO 96/33172 (published Oct. 24, 1996), WO 96/27583(published Mar. 7, 1996), European Patent Application No. 97304971.1(filed Jul. 8, 1997), European Patent Application No. 99308617.2 (filedOct. 29, 1999), WO 98/07697 (published Feb. 26, 1998), WO 98/03516(published Jan. 29, 1998), WO 98/34918 (published Aug. 13, 1998), WO98/34915 (published Aug. 13, 1998), WO 98/33768 (published Aug. 6,1998), WO 98/30566 (published Jul. 16, 1998), European PatentPublication 606,046 (published Jul. 13, 1994), European PatentPublication 931, 788 (published Jul. 28, 1999), WO 90/05719 (publishedMay 31, 1990), WO 99/52910 (published Oct. 21, 1999), WO 99/52889(published Oct. 21, 1999), WO 99/29667 (published Jun. 17, 1999), PCTInternational Application No. PCT/IB98/01113 (filed Jul. 21, 1998),European Patent Application No. 99302232.1 (filed Mar. 25, 1999), GreatBritain Patent Application No. 9912961.1 (filed Jun. 3, 1999), U.S.Provisional Application No. 60/148,464 (filed Aug. 12, 1999), U.S. Pat.No. 5,863,949 (issued Jan. 26, 1999), U.S. Pat. No. 5,861,510 (issuedJan. 19, 1999), and European Patent Publication 780,386 (published Jun.25, 1997), all of which are incorporated herein in their entireties byreference. In some embodiments, MMP-2 and MMP-9 inhibitors are thosethat have little or no activity inhibiting MMP-1. Other embodimentsinclude those that selectively inhibit MMP-2 and/or AMP-9 relative tothe other matrix-metalloproteinases (e.g., MAP-1, MMP-3, MMP-4, MMP-5,MMP-6, MMP-7, MMP-8, MMP-10, MMP-11, MMP-12, and MMP-13). Somenon-limiting examples of MMP inhibitors are AG-3340, RO 32-3555, and RS13-0830.

Autophagy inhibitors include, but are not limited to, chloroquine,3-methyladenine, hydroxychloroquine (Plaquenil™), bafilomycin A1,5-amino-4-imidazole carboxamide riboside (AICAR), okadaic acid,autophagy-suppressive algal toxins which inhibit protein phosphatases oftype 2A or type 1, analogues of cAMP, and drugs which elevate cAMPlevels such as adenosine, LY204002, N6-mercaptopurine riboside, andvinblastine. In addition, antisense or siRNAs that inhibit expression ofproteins including, but not limited to ATG5 (which are implicated inautophagy), can also be used.

In some embodiments, provided herein is a method of and/or apharmaceutical composition for treating a cardiovascular disease in asubject which comprises an amount of a compound provided herein, or apharmaceutically acceptable form (e.g., pharmaceutically acceptablesalts, hydrates, solvates, isomers, prodrugs, and isotopically labeledderivatives) thereof, and an amount of one or more therapeutic agentsuse for the treatment of cardiovascular diseases.

Exemplary agents for use in cardiovascular disease applications areanti-thrombotic agents, e.g., prostacyclin and salicylates, thrombolyticagents, e.g., streptokinase, urokinase, tissue plasminogen activator(TPA) and anisoylated plasminogen-streptokinase activator complex(APSAC), anti-platelets agents, e.g., acetyl-salicylic acid (ASA) andclopidrogel, vasodilating agents, e.g., nitrates, calcium channelblocking drugs, anti-proliferative agents, e.g., colchicine andalkylating agents, intercalating agents, growth modulating factors suchas interleukins, transformation growth factor-beta and congeners ofplatelet derived growth factor, monoclonal antibodies directed againstgrowth factors, anti-inflammatory agents, both steroidal andnon-steroidal, and other agents that can modulate vessel tone, function,arteriosclerosis, and the healing response to vessel or organ injurypost intervention. Antibiotics can also be included in combinations orcoatings. Moreover, a coating can be used to effect therapeutic deliveryfocally within the vessel wall. By incorporation of the active agent ina swellable polymer, the active agent will be released upon swelling ofthe polymer.

In one embodiment, a compound provided herein, or a pharmaceuticallyacceptable form (e.g., pharmaceutically acceptable salts, hydrates,solvates, isomers, prodrugs, and isotopically labeled derivatives)thereof, or a pharmaceutical composition as provided herein, can beformulated or administered in conjunction with liquid or solid tissuebarriers also known as lubricants. Examples of tissue barriers include,but are not limited to, polysaccharides, polyglycans, seprafilm,interceed and hyaluronic acid.

Medicaments which can be administered in conjunction with a compoundprovided herein, or a pharmaceutically acceptable form (e.g.,pharmaceutically acceptable salts, hydrates, solvates, isomers,prodrugs, and isotopically labeled derivatives) thereof, include anysuitable drugs usefully delivered by inhalation for example, analgesics,e.g., codeine, dihydromorphine, ergotamine, fentanyl or morphine;anginal preparations, e.g., diltiazem; antiallergics, e.g. cromoglycate,ketotifen or nedocromil; anti-infectives, e.g., cephalosporins,penicillins, streptomycin, sulphonamides, tetracyclines or pentamidine;antihistamines, e.g., methapyrilene; anti-inflammatories, e.g.,beclomethasone, flunisolide, budesonide, tipredane, triamcinoloneacetonide or fluticasone; antitussives, e.g., noscapine;bronchodilators, e.g., ephedrine, adrenaline, fenoterol, formoterol,isoprenaline, metaproterenol, phenylephrine, phenylpropanolamine,pirbuterol, reproterol, rimiterol, salbutamol, salmeterol, terbutalin,isoetharine, tulobuterol, orciprenaline or(−)-4-amino-3,5-dichloro-α-[[[6-[2-(2-pyridinyl)ethoxy]hexyl]-amino]methyl]benzenemethanol;diuretics, e.g., amiloride; anticholinergics e.g., ipratropium, atropineor oxitropium; hormones, e.g., cortisone, hydrocortisone orprednisolone; xanthines e.g., aminophylline, choline theophyllinate,lysine theophyllinate or theophylline; and therapeutic proteins andpeptides, e.g., insulin or glucagon. It will be clear to a personskilled in the art that, where appropriate, the medicaments can be usedin the form of salts (e.g., as alkali metal or amine salts or as acidaddition salts) or as esters (e.g., lower alkyl esters) to optimize theactivity and/or stability of the medicament.

Other exemplary therapeutic agents useful for a combination therapyinclude, but are not limited to, agents as described above, radiationtherapy, hormone antagonists, hormones and their releasing factors,thyroid and antithyroid drugs, estrogens and progestins, androgens,adrenocorticotropic hormone; adrenocortical steroids and their syntheticanalogs; inhibitors of the synthesis and actions of adrenocorticalhormones, insulin, oral hypoglycemic agents, and the pharmacology of theendocrine pancreas, agents affecting calcification and bone turnover:calcium, phosphate, parathyroid hormone, vitamin D, calcitonin, vitaminssuch as water-soluble vitamins, vitamin B complex, ascorbic acid,fat-soluble vitamins, vitamins A, K, and E, growth factors, cytokines,chemokines, muscarinic receptor agonists and antagonists;anticholinesterase agents; agents acting at the neuromuscular junctionand/or autonomic ganglia; catecholamines, sympathomimetic drugs, andadrenergic receptor agonists or antagonists; and 5-hydroxytryptamine(5-HT, serotonin) receptor agonists and antagonists.

Therapeutic agents can also include agents for pain and inflammationsuch as histamine and histamine antagonists, bradykinin and bradykininantagonists, 5-hydroxytryptamine (serotonin), lipid substances that aregenerated by biotransformation of the products of the selectivehydrolysis of membrane phospholipids, eicosanoids, prostaglandins,thromboxanes, leukotrienes, aspirin, nonsteroidal anti-inflammatoryagents, analgesic-antipyretic agents, agents that inhibit the synthesisof prostaglandins and thromboxanes, selective inhibitors of theinducible cyclooxygenase, selective inhibitors of the induciblecyclooxygenase-2, autacoids, paracrine hormones, somatostatin, gastrin,cytokines that mediate interactions involved in humoral and cellularimmune responses, lipid-derived autacoids, eicosanoids, (3-adrenergicagonists, ipratropium, glucocorticoids, methylxanthines, sodium channelblockers, opioid receptor agonists, calcium channel blockers, membranestabilizers and leukotriene inhibitors.

Additional therapeutic agents contemplated herein include diuretics,vasopressin, agents affecting the renal conservation of water, rennin,angiotensin, agents useful in the treatment of myocardial ischemia,anti-hypertensive agents, angiotensin converting enzyme inhibitors,β-adrenergic receptor antagonists, agents for the treatment ofhypercholesterolemia, and agents for the treatment of dyslipidemia.

Other therapeutic agents contemplated herein include drugs used forcontrol of gastric acidity, agents for the treatment of peptic ulcers,agents for the treatment of gastroesophageal reflux disease, prokineticagents, antiemetics, agents used in irritable bowel syndrome, agentsused for diarrhea, agents used for constipation, agents used forinflammatory bowel disease, agents used for biliary disease, agents usedfor pancreatic disease. Therapeutic agents include, but are not limitedto, those used to treat protozoan infections, drugs used to treatMalaria, Amebiasis, Giardiasis, Trichomoniasis, Trypanosomiasis, and/orLeishmaniasis, and/or drugs used in the chemotherapy of helminthiasis.Other therapeutic agents include, but are not limited to, antimicrobialagents, sulfonamides, trimethoprim-sulfamethoxazole quinolones, andagents for urinary tract infections, penicillins, cephalosporins, andother, β-Lactam antibiotics, an agent containing an aminoglycoside,protein synthesis inhibitors, drugs used in the chemotherapy oftuberculosis, Mycobacterium avium complex disease, and leprosy,antifungal agents, antiviral agents including nonretroviral agents andantiretroviral agents.

Examples of therapeutic antibodies that can be combined with a compoundprovided herein include but are not limited to anti-receptor tyrosinekinase antibodies (cetuximab, panitumumab, trastuzumab), anti CD20antibodies (rituximab, tositumomab), and other antibodies such asalemtuzumab, bevacizumab, and gemtuzumab.

Moreover, therapeutic agents used for immuno-modulation, such asimmuno-modulators, immuno-suppressive agents, tolerogens, andimmunostimulants are contemplated by the methods herein. In addition,therapeutic agents acting on the blood and the blood-forming organs,hematopoietic agents, growth factors, minerals, and vitamins,anticoagulant, thrombolytic, and anti-platelet drugs are alsocontemplated by the methods herein.

In exemplary embodiments, for treating renal carcinoma, one can combinea compound provided herein, or a pharmaceutically acceptable form (e.g.,pharmaceutically acceptable salts, hydrates, solvates, isomers,prodrugs, and isotopically labeled derivatives) thereof, or apharmaceutical composition as provided herein, with sorafenib and/oravastin. For treating an endometrial disorder, one can combine acompound provided herein with doxorubincin, taxotere (taxol), and/orcisplatin (carboplatin). For treating ovarian cancer, one can combine acompound provided herein with cisplatin, carboplatin, docetaxel,doxorubincin, topotecan, and/or tamoxifen. For treating breast cancer,one can combine a compound provided herein with paclitaxel or docetaxel,gemcitabine, capecitabine, tamoxifen, letrozole, erlotinib, lapatinib,PD0325901, bevacizumab, trastuzumab, OSI-906, and/or OSI-930. Fortreating lung cancer, one can combine a compound as provided herein withpaclitaxel, docetaxel, gemcitabine, cisplatin, pemetrexed, erlotinib,PD0325901, and/or bevacizumab.

In some embodiments, the disorder to be treated, prevented and/ormanaged is a hematological cancer, e.g., lymphoma (e.g., T-celllymphoma; NHL), myeloma (e.g., multiple myeloma), and leukemia (e.g.,CLL), and a compound provided herein is used in combination with: HDACinhibitors such as vorinostat, romidepsin and ACY-1215; mTOR inhibitorssuch as everolimus; anti-folates such as pralatrexate; nitrogen mustardsuch as bendamustine; gemcitabine, optionally in further combinationwith oxaliplatin; rituximab-cyclophosphamide combination; PI3Kinhibitors such as RP-6530, TG 100-115, RV1729, GS-1101, XL 499,GDC-0941, and AMG-319; angiogenesis inhibitors such as pomalidomide orBTK inhibitors such as ibrutinib, AVL-292, Dasatinib, LFM-AI3,ONO-WG-307, and GDC-0834. In some embodiments, the disorder to betreated, prevented and/or managed is DLBCL, and a compound providedherein (e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37,38, 40, 41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or apharmaceutically acceptable derivative (e.g., salt or solvate) thereof,is used in combination with HDAC inhibitors provided herein. In oneparticular embodiment, the HDAC inhibitor is ACY-1215.

In some embodiments, the disorder to be treated, prevented and/ormanaged is DLBCL, and a compound provided herein (e.g., compounds 2, 4,7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63,73, 75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptablederivative (e.g., salt or solvate) thereof, is used in combination withBTK inhibitors provided herein. In one particular embodiment, the BTKinhibitor is ibrutinib. In one embodiment, the BTK inhibitor is AVL-292.

In some embodiments, the disorder to be treated, prevented and/ormanaged is DLBCL, and a compound provided herein (e.g., compounds 2, 4,7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63,73, 75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptablederivative (e.g., salt or solvate) thereof, is used in combination withIRAK inhibitors provided herein. In one particular embodiment, the IRAK4inhibitor is ND-2110 or ND-2158.

In some embodiments, the disorder to be treated, prevented and/ormanaged is WM, and a compound provided herein (e.g., compounds 2, 4, 7,9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73,75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptable derivative(e.g., salt or solvate) thereof, is used in combination with BTKinhibitors provided herein. In one particular embodiment, the BTKinhibitor is ibrutinib. In one embodiment, the BTK inhibitor is AVL-292.

In some embodiments, the disorder to be treated, prevented and/ormanaged is WM, and a compound provided herein (e.g., compounds 2, 4, 7,9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73,75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptable derivative(e.g., salt or solvate) thereof, is used in combination with IRAK4inhibitors provided herein. In one particular embodiment, the IRAK4inhibitor is ND-2110 or ND-2158.

In some embodiments, the disorder to be treated, prevented and/ormanaged is T-ALL, the subject/patient has a PTEN deficiency, and acompound provided herein (e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26,27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81,and 88), or a pharmaceutically acceptable derivative (e.g., salt orsolvate) thereof, is used in combination with doxorubicin and/orvincristine.

In certain embodiments, wherein inflammation (e.g., arthritis, asthma)is treated, prevented and/or managed, a compound provided herein can becombined with, for example: PI3K inhibitors such as RP-6530, TG 100-115,RV1729, GS-1101, XL 499, GDC-0941, and AMG-319; BTK inhibitors such asibrutinib and AVL-292; JAK inhibitors such as tofacitinib and GLPG0636;SYK inhibitors such as fostamatinib.

In certain embodiments wherein asthma is treated, prevented and/ormanaged, a compound provided herein can be combined with, for example:beta 2-agonists such as, but not limited to, albuterol (Proventil®, orVentolin®), salmeterol (Serevent®), formoterol (Foradil®),metaproterenol (Alupent®), pirbuterol (MaxAir®), and terbutalinesulfate; corticosteroids such as, but not limited to, budesonide (e.g.,Pulmicort®), flunisolide (e.g., AeroBid Oral Aerosol Inhaler® orNasalide Nasal Aerosol®), fluticasone (e.g., Flonase® or Flovent®) andtriamcinolone (e.g., Azmacort®); mast cell stabilizers such as cromolynsodium (e.g., Intal® or Nasalcrom®) and nedocromil (e.g., Tilade®);xanthine derivatives such as, but not limited to, theophylline (e.g.,Aminophyllin®, Theo-24® or Theolair®); leukotriene receptor antagonistssuch as, but are not limited to, zafirlukast (Accolate®), montelukast(Singulair®), and zileuton (Zyflo®); and adrenergic agonists such as,but are not limited to, epinephrine (Adrenalin®, Bronitin®, EpiPen® orPrimatene Mist®).

In certain embodiments wherein arthritis is treated, prevented and/ormanaged, a compound provided herein can be combined with, for example:TNF antagonist (e.g., a TNF antibody or fragment, a soluble TNF receptoror fragment, fusion proteins thereof, or a small molecule TNFantagonist); other biologic antirhheumatics (e.g., IL-6 antagonists,IL-1 antagonists, costimulatory modulators); an antirheumatic (e.g.,methotrexate, auranofin, aurothioglucose, azathioprine, etanercept, goldsodium thiomalate, chrloroquine, hydroxychloroquine sulfate,leflunomide, sulfasalzine, penicillamine); a muscle relaxant; anarcotic; a non-steroid anti-inflammatory drug (NSAID); an analgesic; ananesthetic; a sedative; a local anesthetic; a neuromuscular blocker; anantimicrobial (e.g., an aminoglycoside, an antifungal, an antiparasitic,an antiviral, a carbapenem, cephalosporin, a fluoroquinolone, amacrolide, a penicillin, a sulfonamide, a tetracycline, anotherantimicrobial); an antipsoriatic; a corticosteroid; an anabolic steroid;a cytokine or a cytokine antagonist; a calcineurin inhibitor (e.g.,cyclosporine, tacrolimus).

In some embodiments, a compound provided herein (e.g., a compound ofFormula I (e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35,37, 38, 40, 41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or anenantiomer or a mixture of enantiomers thereof, or a pharmaceuticallyacceptable salt, solvate, hydrate, co-crystal, clathrate, or polymorphthereof) is administered in combination with an agent for the treatmentof rheumatoid arthritis. Examples of agents for the treatment ofrheumatoid arthritis include, but are not limited to, various NSAIDs,corticosteroids, sulfasalazine, auranofin, methotrexate, azathioprine,penicillamine, cyclosporine, Arava (leflunomide), TNF inhibitors (e.g.,Enbrel (etanercept), Remicade (infliximab), Humira (adalimumab), Simponi(golimumab), and Cimzia (certolizumab)), IL-1 inhibitors (e.g., Kineret(anakinra)), T-cell costimulatory modulators (e.g., Orencia(abatacept)), Anti-CD20 (e.g., Rituxan (rituximab)), and IL-6 inhibitors(e.g., Actemra (tocilizumab)). In one embodiment, the agent is Cimzia(certolizumab). In another embodiment, the agent is Actemra(tocilizumab).

In some embodiments, a compound provided herein (e.g., a compound ofFormula I (e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35,37, 38, 40, 41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or anenantiomer or a mixture of enantiomers thereof, or a pharmaceuticallyacceptable salt, solvate, hydrate, co-crystal, clathrate, or polymorphthereof) is administered in combination with an agent for rheumatology.Examples of agents for rheumatology include, but are not limited to,Rayos (prednisone), Stendra (avanafil), Actemra (tocilizumab), Duexis(ibuprofen and famotidine), Actemra (tocilizumab), Krystexxa(pegloticase), Vimovo (naproxen+esomeprazole), Cimzia (certolizumabpegol), Colcrys (colchicine), Pennsaid (diclofenac sodium topicalsolution), Simponi (golimumab), Uloric (febuxostat), Orencia(abatacept), Elaprase (idursulfase), Orencia (abatacept), Vioxx(rofecoxib), Enbrel (etanercept), Humira (adalimumab), Remicade(infliximab), Bextra, Kineret, Remicade (infliximab), Supartz, Mobic(meloxicam), Vivelle (estradiol transdermal system), Lodine XL(etodolac), Arava, Salagen, Arthrotec, Etodolac, Ketoprofen, Synvisc,Tolmetin Sodium, Azulfidine EN-tabs Tablets (sulfasalazine delayedrelease tablets, USP), and Naprelan (naproxen sodium).

In some embodiments, the second agent is selected from belimumab,AGS-009, rontalizumab, vitamin D3, sifalimumab, AMG 811, IFNα Kinoid,CEP33457, epratuzumab, LY2127399, Ocrelizumab, Atacicept, A-623,SBI-087, AMG557, laquinimod, rapamycin, cyclophosphamide, azathioprine,mycophenolate, leflunomide, methotrexate, CNTO 136, tamibarotene,N-acetylcysteine, CDP7657, hydroxychloroquine, rituximab, carfilzomib,bortezomib, ONX 0914, IMO-3100, DV1179, sulfasalazine, and chloroquine.In one embodiment, the second agent is methotrexate, sulfasalazine,chloroquine, or hydroxychloroquine. In one embodiment, the second agentis methotrexate.

In certain embodiments wherein psoriasis is treated, prevented and/ormanaged, a compound provided herein can be combined with, for example:budesonide, epidermal growth factor, corticosteroids, cyclosporine,sulfasalazine, aminosalicylates, 6-mercaptopurine, azathioprine,metronidazole, lipoxygenase inhibitors, mesalamine, olsalazine,balsalazide, antioxidants, thromboxane inhibitors, IL-1 receptorantagonists, anti-IL-1β monoclonal antibodies, anti-IL-6 monoclonalantibodies, growth factors, elastase inhibitors, pyridinyl-imidazolecompounds, antibodies or agonists of TNF, LT, IL-1, IL-2, IL-6, IL-7,IL-8, IL-15, IL-16, IL-18, EMAP-II, GM-CSF, FGF, and PDGF, antibodies ofCD2, CD3, CD4, CD8, CD25, CD28, CD30, CD40, CD45, CD69, CD90 or theirligands, methotrexate, cyclosporine, FK506, rapamycin, mycophenolatemofetil, leflunomide, NSAIDs, ibuprofen, corticosteroids, prednisolone,phosphodiesterase inhibitors, adenosine agonists, antithrombotic agents,complement inhibitors, adrenergic agents, IRAK, NIK, IKK, p38, MAPkinase inhibitors, IL-1β converting enzyme inhibitors, TNFα convertingenzyme inhibitors, T-cell signaling inhibitors, metalloproteinaseinhibitors, sulfasalazine, azathioprine, 6-mercaptopurines, angiotensinconverting enzyme inhibitors, soluble cytokine receptors, soluble p55TNF receptor, soluble p75 TNF receptor, sIL-1RI, sIL-1RII, sIL-6R,anti-inflammatory cytokines, IL-4, IL-10, IL-11, IL-13 and TGFβ.

In certain embodiments wherein fibrosis or fibrotic condition of thebone marrow is treated, prevented and/or managed, a compound providedherein can be combined with, for example, a Jak2 inhibitor (including,but not limited to, INCB018424, XL019, TG101348, or TG101209), animmuno-modulator, e.g., an IMID® (including, but not limited tothalidomide, lenalidomide, or panolinomide), hydroxyurea, an androgen,erythropoietic stimulating agents, prednisone, danazol, HDAC inhibitors,or other agents or therapeutic modalities (e.g., stem cell transplants,or radiation).

In certain embodiments wherein fibrosis or fibrotic condition of theheart is treated, prevented and/or managed, a compound provided hereincan be combined with, for example, eplerenone, furosemide, pycnogenol,spironolactone, TcNC100692, torasemide (e.g., prolonged release form oftorasemide), or combinations thereof.

In certain embodiments wherein fibrosis or fibrotic condition of thekidney is treated, prevented and/or managed, a compound provided hereincan be combined with, for example, cyclosporine, cyclosporine A,daclizumab, everolimus, gadofoveset trisodium (ABLAVAR®), imatinibmesylate (GLEEVEC®), matinib mesylate, methotrexate, mycophenolatemofetil, prednisone, sirolimus, spironolactone, STX-100, tamoxifen,TheraCLEC™, or combinations thereof.

In certain embodiments wherein fibrosis or fibrotic condition of theskin is treated, prevented and/or managed, a compound provided hereincan be combined with, for example, Bosentan (Tracleer), p144,pentoxifylline; pirfenidone; pravastatin, STI571, Vitamin E, orcombinations thereof.

In certain embodiments wherein fibrosis or fibrotic condition of thegastrointestinal system is treated, prevented and/or managed, a compoundprovided herein can be combined with, for example, ALTU-135, bucelipasealfa (INN), DCI1020, EUR-1008 (ZENPEP™), ibuprofen, Lym-X-Sorb powder,pancrease MT, pancrelipase (e.g., pancrelipase delayed release), pentadecanoic acid (PA), repaglinide, TheraCLEC™, triheptadecanoin (THA),ULTRASE MT20, ursodiol, or combinations thereof.

In certain embodiments wherein fibrosis or fibrotic condition of thelung is treated, prevented and/or managed, a compound provided hereincan be combined with, for example, 18-FDG, AB0024, ACT-064992(macitentan), aerosol interferon-gamma, aerosolized human plasma-derivedalpha-1 antitrypsin, alpha1-proteinase inhibitor, ambrisentan, amikacin,amiloride, amitriptyline, anti-pseudomonas IgY gargle, ARIKACE™,AUREXIS® (tefibazumab), AZAPRED, azathioprine, azithromycin,azithromycin, AZLI, aztreonam lysine, BIBF1120, Bio-25 probiotic,bosentan, Bramitob®, calfactant aerosol, captopril, CC-930, ceftazidime,ceftazidime, cholecalciferol (Vitamin D3), ciprofloxacin (CIPRO®,BAYQ3939), CNTO 888, colistin CF, combined Plasma Exchange (PEX),rituximab, and corticosteroids, cyclophosphamide, dapsone, dasatinib,denufosol tetrasodium (INS37217), dornase alfa (PULMOZYME®), EPI-hNE4,erythromycin, etanercept, FG-3019, fluticasone, FTI, GC1008, GS-9411,hypertonic saline, ibuprofen, iloprost inhalation, imatinib mesylate(GLEEVEC®), inhaled sodium bicarbonate, inhaled sodium pyruvate,interferon gamma-1b, interferon-alpha lozenges, isotonic saline, IW001,KB001, losartan, lucinactant, mannitol, meropenem, meropenem infusion,miglustat, minocycline, Moli1901, MP-376 (levofloxacin solution forinhalation), mucoid exopolysaccharide P. aeruginosa immune globulin IV,mycophenolate mofetil, n-acetylcysteine, N-acetylcysteine (NAC), NaCl6%, nitric oxide for inhalation, obramycin, octreotide, oligoG CF-5/20,Omalizumab, pioglitazone, piperacillin-tazobactam, pirfenidone,pomalidomide (CC-4047), prednisone, prevastatin, PRM-151, QAX576,rhDNAse, SB656933, SB-656933-AAA, sildenafil, tamoxifen, technetium[Tc-99m] sulfur colloid and Indium [In-111] DTPA, tetrathiomolybdate,thalidomide, ticarcillin-clavulanate, tiotropium bromide, tiotropiumRESPIMAT® inhaler, tobramycin (GERNEBCIN®), treprostinil, uridine,valganciclovir (VALCYTE®), vardenafil, vitamin D3, xylitol, zileuton, orcombinations thereof.

In certain embodiments wherein fibrosis or fibrotic condition of theliver is treated, prevented and/or managed, a compound provided hereincan be combined with, for example, adefovir dipivoxil, candesartan,colchicine, combined ATG, mycophenolate mofetil, and tacrolimus,combined cyclosporine microemulsion and tacrolimus, elastometry,everolimus, FG-3019, Fuzheng Huayu, GI262570, glycyrrhizin (monoammoniumglycyrrhizinate, glycine, L-cysteine monohydrochloride), interferongamma-1b, irbesartan, losartan, oltipraz, ORAL IMPACT®, peginterferonalfa-2a, combined peginterferon alfa-2a and ribavirin, peginterferonalfa-2b (SCH 54031), combined peginterferon alpha-2b and ribavirin,praziquantel, prazosin, raltegravir, ribavirin (REBETOL®, SCH 18908),ritonavir-boosted protease inhibitor, pentoxyphilline, tacrolimus,tauroursodeoxycholic acid, tocopherol, ursodiol, warfarin, orcombinations thereof.

In certain embodiments wherein cystic fibrosis is treated, preventedand/or managed, a compound provided herein can be combined with, forexample, 552-02, 5-methyltetrahydrofolate and vitamin B12, Ad5-CB-CFTR,Adeno-associated virus-CFTR vector, albuterol, alendronate, alphatocopherol plus ascorbic acid, amiloride HCl, aquADEK™, ataluren(PTC124), AZD1236, AZD9668, azithromycin, bevacizumab, biaxin(clarithromycin), BIIL 283 BS (amelubent), buprofen, calcium carbonate,ceftazidime, cholecalciferol, choline supplementation, CPX, cysticfibrosis transmembrane conductance regulator, DHA-rich supplement,digitoxin, cocosahexaenoic acid (DHA), doxycycline, ECGC, ecombinanthuman IGF-1, educed glutathione sodium salt, ergocalciferol (vitaminD2), fluorometholone, gadobutrol (GADOVIST®, BAY86-4875), gentamicin,ghrelin, glargine, glutamine, growth hormone, GS-9411, H5.001CBCFTR,human recombinant growth hormone, hydroxychloroquine, hyperbaric oxygen,hypertonic saline, IH636 grape seed proanthocyanidin extract, insulin,interferon gamma-1b, IoGen (molecular iodine), iosartan potassium,isotonic saline, itraconazole, IV gallium nitrate (GANITE®) infusion,ketorolac acetate, lansoprazole, L-arginine, linezolid, lubiprostone,meropenem, miglustat, MP-376 (levofloxacin solution for inhalation),normal saline IV, Nutropin AQ, omega-3 triglycerides, pGM169/GL67A,pGT-1 gene lipid complex, pioglitazone, PTC124, QAU145, salmeterol,SB656933, SB656933, simvastatin, sitagliptin, sodium 4-phenylbutyrate,standardized turmeric root extract, tgAAVCF, TNF blocker, TOBI,tobramycin, tocotrienol, unconjugated Isoflavones 100, vitamin: cholinebitartrate (2-hydroxyethyl) trimethylammonium salt 1:1, VX-770, VX-809,Zinc acetate, or combinations thereof.

In some embodiments, a compound provided herein is administered incombination with an agent that inhibits IgE production or activity. Insome embodiments, the PI3K inhibitor (e.g., PI3Kδ inhibitor) isadministered in combination with an inhibitor of mTOR. Agents thatinhibit IgE production are known in the art and they include but are notlimited to one or more of TEI-9874,2-(4-(6-cyclohexyloxy-2-naphtyloxy)phenylacetamide)benzoic acid,rapamycin, rapamycin analogs (i.e. rapalogs), TORC1 inhibitors, TORC2inhibitors, and any other compounds that inhibit mTORC1 and mTORC2.Agents that inhibit IgE activity include, for example, anti-IgEantibodies such as for example Omalizumab and TNX-901.

In certain embodiments wherein scleroderma is treated, prevented and/ormanaged, a compound provided herein can be combined with, for example:an immunosuppressant (e.g., methotrexate, azathioprine (Imuran®),cyclosporine, mycophenolate mofetil (Cellcept®), and cyclophosphamide(Cytoxan®)); T-cell-directed therapy (e.g., halofuginone, basiliximab,alemtuzumab, abatacept, rapamycin); B-cell directed therapy (e.g.,rituximab); autologous hematopoietic stem cell transplantation; achemokine ligand receptor antagonist (e.g., an agent that targets theCXCL12/CSCR4 axis (e.g., AMD3100)); a DNA methylation inhibitor (e.g.,5-azacytidine); a histone deacetylase inhibitor (e.g., trichostatin A);a statin (e.g., atorvastatin, simvastatin, pravastatin); an endothelinreceptor antagonist (e.g., Bosentan®); a phosphodiesterase type Vinhibitor (e.g., Sildenafil®); a prostacyclin analog (e.g.,trepostinil); an inhibitor of cytokine synthesis and/or signaling (e.g.,Imatinib mesylate, Rosiglitazone, rapamycin, antitransforming growthfactor β1 (anti-TGFβ1) antibody, mycophenolate mofetil, an anti-IL-6antibody (e.g., tocilizumab)); corticosteroids; nonsteroidalanti-inflammatory drugs; light therapy; and blood pressure medications(e.g., ACE inhibitors).

In certain embodiments wherein inflammatory myopathies are treated,prevented and/or managed, a compound provided herein can be combinedwith, for example: topical creams or ointments (e.g., topicalcorticosteroids, tacrolimus, pimecrolimus); cyclosporine (e.g., topicalcyclosporine); an anti-interferon therapy, e.g., AGS-009, Rontalizumab(rhuMAb IFNalpha), Vitamin D3, Sifalimumab (MEDI-545), AMG 811, IFNαKinoid, or CEP33457. In some embodiments, the other therapy is an IFN-αtherapy, e.g., AGS-009, Rontalizumab, Vitamin D3, Sifalimumab (MEDI-545)or IFNα Kinoid; corticosteroids such as prednisone (e.g., oralprednisone); immunosuppressive therapies such as methotrexate (Trexall®,Methotrexate®, Rheumatrex®), azathioprine (Azasan®, Imuran®),intravenous immunoglobulin, tacrolimus (Prograf®), pimecrolimus,cyclophosphamide (Cytoxan®), and cyclosporine (Gengraf®, Neoral®,Sandimmune®); anti-malarial agents such as hydroxychloroquine(Plaquenil®) and chloroquine (Aralen®); total body irradiation;rituximab (Rituxan®); TNF inhibitors (e.g., etanercept (Enbrel®),infliximab (Remicade®)); AGS-009; Rontalizumab (rhuMAb IFNalpha);Vitamin D3; Sifalimumab (MEDI-545); AMG 811; IFNα Kinoid; CEP33457;agents that inhibit IgE production such as TEI-9874,2-(4-(6-cyclohexyloxy-2-naphtyloxy)phenylacetamide)benzoic acid,rapamycin, rapamycin analogs (i.e. rapalogs), TORC1 inhibitors, TORC2inhibitors, and any other compounds that inhibit mTORC1 and mTORC2;agents that inhibit IgE activity such as anti-IgE antibodies (e.g.,Omalizumab and TNX-90); and additional therapies such as physicaltherapy, exercise, rest, speech therapy, sun avoidance, heat therapy,and surgery.

In certain embodiments wherein myositis (e.g., dermatomysitis) istreated, prevented and/or managed, a compound provided herein can becombined with, for example: corticosteroids; corticosteroid sparingagents such as, but not limited to, azathioprine and methotrexate;intravenous immunoglobulin; immunosuppressive agents such as, but notlimited to, tacrolimus, cyclophosphamide and cyclosporine; rituximab;TNFα inhibitors such as, but not limited to, etanercept and infliximab;growth hormone; growth hormone secretagogues such as, but not limitedto, MK-0677, L-162752, L-163022, NN703 ipamorelin, hexarelin, GPA-748(KP102, GHRP-2), and LY444711 (Eli Lilly); other growth hormone releasestimulators such as, but not limited to, Geref, GHRH (1-44), Somatorelin(GRF 1-44), ThGRF genotropin, L-DOPA, glucagon, and vasopressin; andinsulin-like growth factor.

In certain embodiments wherein Sjögren's syndrome is treated, preventedand/or managed, a compound provided herein can be combined with, forexample: pilocarpine; cevimeline; nonsteroidal anti-inflammatory drugs;arthritis medications; antifungal agents; cyclosporine;hydroxychloroquine; prednisone; azathioprine; and cyclophamide.

Further therapeutic agents that can be combined with a compound providedherein can be found in Goodman and Gilman's “The Pharmacological Basisof Therapeutics” Tenth Edition edited by Hardman, Limbird and Gilman orthe Physician's Desk Reference, both of which are incorporated herein byreference in their entirety.

In one embodiment, the compounds described herein can be used incombination with the agents provided herein or other suitable agents,depending on the condition being treated. Hence, in some embodiments, acompound provided herein, or a pharmaceutically acceptable form thereof,will be co-administered with other agents as described above. When usedin combination therapy, a compound described herein, or apharmaceutically acceptable form thereof, can be administered with asecond agent simultaneously or separately. This administration incombination can include simultaneous administration of the two agents inthe same dosage form, simultaneous administration in separate dosageforms, and separate administration. That is, a compound described hereinand any of the agents described above can be formulated together in thesame dosage form and administered simultaneously. Alternatively, acompound provided herein and any of the agents described above can besimultaneously administered, wherein both agents are present in separateformulations. In another alternative, a compound provided herein can beadministered just followed by any of the agents described above, or viceversa. In the separate administration protocol, a compound providedherein and any of the agents described above can be administered a fewminutes apart, or a few hours apart, or a few days apart.

Administration of a compound provided herein, or a pharmaceuticallyacceptable form thereof, can be effected by any method that enablesdelivery of the compound to the site of action. An effective amount of acompound provided herein, or a pharmaceutically acceptable form thereof,can be administered in either single or multiple doses by any of theaccepted modes of administration of agents having similar utilities,including rectal, buccal, intranasal, and transdermal routes, byintra-arterial injection, intravenously, intraperitoneally,parenterally, intramuscularly, subcutaneously, orally, topically, as aninhalant, or via an impregnated or coated device such as a stent, forexample, or an artery-inserted cylindrical polymer.

When a compound provided herein, or a pharmaceutically acceptable formthereof, is administered in a pharmaceutical composition that comprisesone or more agents, and the agent has a shorter half-life than thecompound provided herein, unit dose forms of the agent and the compoundas provided herein can be adjusted accordingly.

In some embodiments, the compound provided herein and the second agentare administered as separate compositions, e.g., pharmaceuticalcompositions. In some embodiments, the PI3K modulator and the agent areadministered separately, but via the same route (e.g., both orally orboth intravenously). In other embodiments, the PI3K modulator and theagent are administered in the same composition, e.g., pharmaceuticalcomposition.

In some embodiments, a compound provided herein (e.g., a compound ofFormula I (e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35,37, 38, 40, 41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or anenantiomer or a mixture of enantiomers thereof, or a pharmaceuticallyacceptable salt, solvate, hydrate, co-crystal, clathrate, or polymorphthereof) is administered in combination with an agent for pulmonary orrespiratory diseases. Examples of agents for pulmonary or respiratorydiseases include, but are not limited to, Dymista (azelastinehydrochloride and fluticasone propionate), Kalydeco (ivacaftor), Qnasl(beclomethasone dipropionate) nasal aerosol, Rayos (prednisone)delayed-release tablets, Surfaxin (lucinactant), Tudorza Pressair(aclidinium bromide inhalation powder), Arcapta (indacaterol maleateinhalation powder), Daliresp (roflumilast), Xalkori (crizotinib),Cayston (aztreonam for inhalation solution), Dulera (mometasonefuroate+formoterol fumarate dihydrate), Teflaro (ceftaroline fosamil),Adcirca (tadalafil), Tyvaso (treprostinil), Alvesco (ciclesonide),Patanase (olopatadine hydrochloride), Letairis (ambrisentan), Xyzal(levocetirizine dihydrochloride), Brovana (arformoterol tartrate),Tygacil (tigecycline), Ketek (telithromycin), Spiriva HandiHaler(tiotropium bromide), Aldurazyme (laronidase), Iressa (gefitinib),Xolair (omalizumab), Zemaira (alpha1-proteinase inhibitor), Clarinex,Qvar (beclomethasone dipropionate), Remodulin (treprostinil), Xopenex,Avelox I.V. (moxifloxacin hydrochloride), DuoNeb (albuterol sulfate andipratropium bromide), Foradil Aerolizer (formoterol fumarate inhalationpowder), Invanz, NasalCrom Nasal Spray, Tavist (clemastine fumarate),Tracleer (bosentan), Ventolin HFA (albuterol sulfate inhalationaerosol), Biaxin XL (clarithromycin extended-release tablets), Cefazolinand Dextrose USP, Tri-Nasal Spray (triamcinolone acetonide spray),Accolate, Cafcit Injection, Proventil HFA Inhalation Aerosol, RhinocortAqua Nasal Spray, Tequin, Tikosyn Capsules, Allegra-D, Clemastinefumarate syrup, Curosurf, Dynabac, Infasurf, Priftin, Pulmozyme (dornasealfa), Sclerosol Intrapleural Aerosol, Singulair, Synagis, Ceftin(cefuroxime axetil), Cipro (ciprofloxacin HCl), Claritin RediTabs (10 mgloratadine rapidly-disintegrating tablet), Flonase Nasal Spray, FloventRotadisk, Metaprotereol Sulfate Inhalation Solution (5%), Nasacort AQ(triamcinolone acetonide) Nasal Spray, Omnicef, Raxar (grepafloxacin),Serevent, Tilade (nedocromil sodium), Tobi, Vanceril 84 mcg DoubleStrength (beclomethasone dipropionate, 84 mcg) Inhalation Aerosol, Zagam(sparfloxacin) tablets, Zyflo (Zileuton), Accolate, Allegra(fexofenadine hydrochloride), Astelin nasal spray, Atrovent (ipratropiumbromide), Augmentin (amoxicillin/clavulanate), Azmacort (triamcinoloneacetonide) Inhalation Aerosol, Breathe Right, Claritin Syrup(loratadine), Claritin-D 24 Hour Extended Release Tablets (10 mgloratadine, 240 mg pseudoephedrine sulfate), Covera-HS (verapamil),Nasacort AQ (triamcinolone acetonide) Nasal Spray, OcuHist, Pulmozyme(dornase alfa), RespiGam (Respiratory Syncitial Virus Immune GlobulinIntravenous), Tavist (clemastine fumarate), Tripedia (Diptheria andTetanus Toxoids and Acellular Pertussis Vaccine Absorbed), Vancenase AQ84 mcg Double Strength, Visipaque (iodixanol), Zosyn (sterilepiperacillin sodium/tazobactam sodium), Cedax (ceftibuten), and Zyrtec(cetirizine HCl). In one embodiment, the agent for pulmonary orrespiratory diseases is Arcapta, Daliresp, Dulera, Alvesco, Brovana,Spiriva HandiHaler, Xolair, Qvar, Xopenex, DuoNeb, Foradil Aerolizer,Accolate, Singulair, Flovent Rotadisk, Tilade, Vanceril, Zyflo, orAzmacort Inhalation Aerosol. In one embodiment, the agent for pulmonaryor respiratory diseases is Spiriva HandiHaler.

In some embodiments, a compound provided herein (e.g., a compound ofFormula I (e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35,37, 38, 40, 41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or anenantiomer or a mixture of enantiomers thereof, or a pharmaceuticallyacceptable salt, solvate, hydrate, co-crystal, clathrate, or polymorphthereof) is administered in combination with an agent for immunology orinfectious diseases. Examples of agents for immunology or infectiousdiseases include, but are not limited to, Horizant (gabapentinenacarbil), Qnasl (beclomethasone dipropionate) nasal aerosol, Rayos(prednisone) delayed-release tablets, Stribild (elvitegravir,cobicistat, emtricitabine, tenofovir disoproxil fumarate), TudorzaPressair (aclidinium bromide inhalation powder), Arcapta (indacaterolmaleate inhalation powder), Benlysta (belimumab), Complera(emtricitabine/rilpivirine/tenofovir disoproxil fumarate), Daliresp(roflumilast), Dificid (fidaxomicin), Edurant (rilpivirine), Firazyr(icatibant), Gralise (gabapentin), Incivek (telaprevir), Nulojix(belatacept), Victrelis (boceprevir), Cayston (aztreonam for inhalationsolution), Egrifta (tesamorelin for injection), Menveo (meningitisvaccine), Oravig (miconazole), Prevnar 13 (Pneumococcal 13-valentConjugate Vaccine), Teflaro (ceftaroline fosamil), Zortress(everolimus), Zymaxid (gatifloxacin ophthalmic solution), Bepreve(bepotastine besilate ophthalmic solution), Berinert (C1 EsteraseInhibitor (Human)), Besivance (besifloxacin ophthalmic suspension),Cervarix [Human Papillomavirus Bivalent (Types 16 and 18) Vaccine,Recombinant], Coartem (artemether/lumefantrine), Hiberix (Haemophilus bConjugate Vaccine; Tetanus Toxoid Conjugate), Ilaris (canakinumab),Ixiaro (Japanese Encephalitis Vaccine, Inactivated, Adsorbed), Kalbitor(ecallantide), Qutenza (capsaicin), Vibativ (telavancin), Zirgan(ganciclovir ophthalmic gel), Aptivus (tipranavir), Astepro (azelastinehydrochloride nasal spray), Cinryze (C1 Inhibitor (Human)), Intelence(etravirine), Moxatag (amoxicillin), Rotarix (Rotavirus Vaccine, Live,Oral), Tysabri (natalizumab), Viread (tenofovir disoproxil fumarate),Altabax (retapamulin), AzaSite (azithromycin), Doribax (doripenem),Extina (ketoconazole), Isentress (raltegravir), Selzentry (maraviroc),Veramyst (fluticasone furoate), Xyzal (levocetirizine dihydrochloride),Eraxis (anidulafungin), Gardasil (quadrivalent human papillomavirus(types 6, 11, 16, 18) recombinant vaccine), Noxafil (posaconazole),Prezista (darunavir), Rotateq (rotavirus vaccine, live oralpentavalent), Tyzeka (telbivudine), Veregen (kunecatechins), Aptivus(tipranavir), Baraclude (entecavir), Tygacil (tigecycline), Ketek(telithromycin), Tindamax, tinidazole, Xifaxan (rifaximin), Amevive(alefacept), FluMist (Influenza Virus Vaccine), Fuzeon (enfuvirtide),Lexiva (fosamprenavir calcium), Reyataz (atazanavir sulfate), Alinia(nitazoxanide), Clarinex, Daptacel, Fluzone Preservative-free, Hepsera(adefovir dipivoxil), Pediarix Vaccine, Pegasys (peginterferon alfa-2a),Restasis (cyclosporine ophthalmic emulsion), Sustiva, Vfend(voriconazole), Avelox I.V. (moxifloxacin hydrochloride), Cancidas,Peg-Intron (peginterferon alfa-2b), Rebetol (ribavirin), Spectracef,Twinrix, Valcyte (valganciclovir HCl), Viread (tenofovir disoproxilfumarate), Xigris (drotrecogin alfa [activated]), ABREVA (docosanol),Biaxin XL (clarithromycin extended-release tablets), Cefazolin andDextrose USP, Children's Motrin Cold, Evoxac, Kaletra Capsules and OralSolution, Lamisil (terbinafine hydrochloride) Solution (1%), Lotrisone(clotrimazole/betamethasone diproprionate) lotion, Malarone (atovaquone;proguanil hydrochloride) Tablet, Rapamune (sirolimus) Tablets, RidMousse, Tri-Nasal Spray (triamcinolone acetonide spray), Trivagizole 3(clotrimazole) Vaginal Cream, Trizivir (abacavir sulfate; lamivudine;zidovudine AZT) Tablet, Agenerase (amprenavir), Cleocin (clindamycinphosphate), Famvir (famciclovir), Norvir (ritonavir), Panretin Gel,Rapamune (sirolimus) oral solution, Relenza, Synercid I.V., Tamiflucapsule, Vistide (cidofovir), Allegra-D, CellCept, Clemastine fumaratesyrup, Cleocin (clindamycin phosphate), Dynabac, REBETRON™ CombinationTherapy, Simulect, Timentin, Viroptic, INFANRIX (Diphtheria and TetanusToxoids and Acellular Pertussis Vaccine Adsorbed), Acyclovir Capsules,Aldara (imiquimod), Aphthasol, Combivir, Condylox Gel 0.5% (pokofilox),Famvir (famciclovir), Flagyl ER, Flonase Nasal Spray, Fortovase,INFERGEN (interferon alfacon-1), Intron A (interferon alfa-2b,recombinant), Norvir (ritonavir), Rescriptor Tablets (delavirdinemesylate tablets), SPORANOX (itraconazole), Stromectol (ivermectin),Taxol, Trovan, VIRACEPT (nelfinavir mesylate), Zerit (stavudine),Albenza (albendazole), Apthasol (Amlexanox), Carrington patch, Confide,Crixivan (Indinavir sulfate), Gastrocrom Oral Concentrate (cromolynsodium), Havrix, Lamisil (terbinafine hydrochloride) Tablets, Leukine(sargramostim), Oral Cytovene, RespiGam (Respiratory Syncitial VirusImmune Globulin Intravenous), Videx (didanosine), Viramune (nevirapine),Vistide (cidofovir), Vitrasert Implant, Zithromax (azithromycin), Cedax(ceftibuten), Clarithromycin (Biaxin), Epivir (lamivudine), Intron A(Interferon alfa-2b, recombinant), Invirase (saquinavir), Valtrex(valacyclovir HCl), Western blot confirmatory device, Zerit (stavudine),and Zyrtec (cetirizine HCl).

In some embodiments, the second agent is an HDAC inhibitor, such as,e.g., belinostat, vorinostat, panobinostat, ACY-1215, or romidepsin.

In some embodiments, the second agent is an mTOR inhibitor, such as,e.g., everolimus (RAD 001).

In some embodiments, the second agent is a proteasome inhibitor, suchas, e.g., bortezomib or carfilzomib.

In some embodiments, the second agent is a PKC-β inhibitor, such as,e.g., Enzastaurin (LY317615).

In some embodiments, the second agent is a JAK/STAT inhibitor, such as,e.g., INCB16562 or AZD1480.

In some embodiments, the second agent is an anti-folate, such as, e.g.,pralatrexate.

In some embodiments, the second agent is a farnesyl transferaseinhibitor, such as, e.g., tipifarnib.

In some embodiments, the second agent is an antibody or a biologicagent, such as, e.g., alemtuzumab, rituximab, ofatumumab, or brentuximabvedotin (SGN-035). In one embodiment, the second agent is rituximab. Inone embodiment, the second agent is rituximab and the combinationtherapy is for treating, preventing, and/or managing iNHL, FL, splenicmarginal zone, nodal marginal zone, extranodal marginal zone, and/orSLL.

In some embodiments, a compound provided herein (e.g., compounds 2, 4,7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63,73, 75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptablederivative (e.g., salt or solvate) thereof, is used in combinationbendamustine and one additional active agent. In one embodiment, thecancer or hematological malignancy is iNHL.

In some embodiments, a compound provided herein (e.g., compounds 2, 4,7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63,73, 75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptablederivative (e.g., salt or solvate) thereof, is used in combinationrituximab and one additional active agent. In one embodiment, the canceror hematological malignancy is iNHL.

In some embodiments, a compound provided herein (e.g., compounds 2, 4,7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63,73, 75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptablederivative (e.g., salt or solvate) thereof, is used in combinationbendamustine and rituximab. In one embodiment, the cancer orhematological malignancy is iNHL.

In some embodiments, a compound provided herein (e.g., compounds 2, 4,7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63,73, 75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptablederivative (e.g., salt or solvate) thereof, is used in combinationfludarabine, cyclophosphamide, and rituximab. In one embodiment, thecancer or hematological malignancy is CLL.

In some embodiments, a compound provided herein (e.g., compounds 2, 4,7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63,73, 75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptablederivative (e.g., salt or solvate) thereof, is used in combination withan antibody or a biologic agent, such as, e.g., alemtuzumab, rituximab,ofatumumab, or brentuximab vedotin (SGN-035). In one embodiment, thesecond agent is rituximab. In one embodiment, the second agent isrituximab and the combination therapy is for treating, preventing,and/or managing iNHL, FL, splenic marginal zone, nodal marginal zone,extranodal marginal zone, and/or SLL.

In some embodiments, a compound provided herein (e.g., compounds 2, 4,7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63,73, 75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptablederivative (e.g., salt or solvate) thereof, is used in combination withan antibody-drug conjugate, such as, e.g., inotuzumab ozogamicin, orbrentuximab vedotin.

In some embodiments, a compound provided herein (e.g., compounds 2, 4,7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63,73, 75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptablederivative (e.g., salt or solvate) thereof, is used in combination witha cytotoxic agent, such as, e.g., bendamustine, gemcitabine,oxaliplatin, cyclophosphamide, vincristine, vinblastine, anthracycline(e.g., daunorubicin or daunomycin, doxorubicin), actinomycin,dactinomycin, bleomycin, clofarabine, nelarabine, cladribine,asparaginase, methotrexate, or pralatrexate.

In some embodiments, a compound provided herein (e.g., compounds 2, 4,7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63,73, 75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptablederivative (e.g., salt or solvate) thereof, is used in combination withone or more other anti-cancer agents or chemotherapeutic agents, suchas, e.g., fludarabine, ibrutinib, fostamatinib, lenalidomide,thalidomide, rituximab, cyclophosphamide, doxorubicin, vincristine,prednisone, or R-CHOP (Rituximab, Cyclophosphamide, Doxorubicin orHydroxydaunomycin, Vincristine or Oncovin, Prednisone).

In some embodiments, a compound provided herein (e.g., compounds 2, 4,7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63,73, 75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptablederivative (e.g., salt or solvate) thereof, is used in combination withan antibody for a cytokine (e.g., an IL-15 antibody, an IL-21 antibody,an IL-4 antibody, an IL-7 antibody, an IL-2 antibody, an IL-9 antibody).In some embodiments, the second agent is a JAK1 inhibitor, a JAK3inhibitor, a pan-JAK inhibitor, a BTK inhibitor, an SYK inhibitor, or aPI3K delta inhibitor. In some embodiments, the second agent is anantibody for a chemokine.

Without being limited to a particular theory, a targeted combinationtherapy described herein has reduced side effect and/or enhancedefficacy. For example, in one embodiment, provided herein is acombination therapy for treating CLL with a compound described herein(e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40,41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or apharmaceutically acceptable derivative (e.g., salt or solvate) thereof,and a second active agent (e.g., IL-15 antibodies, IL-21 antibodies,IL-4 antibodies, IL-7 antibodies, IL-2 antibodies, IL-9 antibodies, JAK1inhibitors, JAK3 inhibitors, pan-JAK inhibitors, BTK inhibitors, SYKinhibitors, and/or PI3K delta inhibitors).

Further without being limited by a particular theory, it was found thata compound provided herein (e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26,27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81,and 88) does not affect BTK or MEK pathway. Accordingly, in someembodiments, provided herein is a method of treating or managing canceror hematological malignancy comprising administering to a patient atherapeutically effective amount of a compound provided herein (e.g.,compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41,52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or a pharmaceuticallyacceptable derivative (e.g., salt or solvate) thereof, in combinationwith a BTK inhibitor. In one embodiment, the BTK inhibitor is ibrutinib.In one embodiment, the BTK inhibitor is AVL-292. In one embodiment, thecancer or hematological malignancy is DLBCL. In another embodiment, thecancer or hematological malignancy is iNHL. In another embodiment, thecancer or hematological malignancy is CLL.

In other embodiments, provided herein is a method of treating ormanaging cancer or hematological malignancy comprising administering toa patient a therapeutically effective amount of a compound providedherein (e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37,38, 40, 41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or apharmaceutically acceptable derivative (e.g., salt or solvate) thereof,in combination with a MEK inhibitor. In one embodiment, the MEKinhibitor is trametinib/GSK1120212(N-(3-{3-Cyclopropyl-5-[(2-fluoro-4-iodophenyl)amino]-6,8-dimethyl-2,4,7-trioxo-3,4,6,7-tetrahydropyrido[4,3-d]pyrimidin-1(2H)-yl}phenyl)acetamide),selumetinob(6-(4-bromo-2-chloroanilino)-7-fluoro-N-(2-hydroxyethoxy)-3-methylbenzimidazole-5-carboxamide),pimasertib/AS703026/MSC1935369((S)—N-(2,3-dihydroxypropyl)-3-((2-fluoro-4-iodophenyl)amino)isonicotinamide),XL-518/GDC-0973(1-({3,4-difluoro-2-[(2-fluoro-4-iodophenyl)amino]phenyl}carbonyl)-3-[(2S)-piperidin-2-yl]azetidin-3-ol),refametinib/BAY869766/RDEA119(N-(3,4-difluoro-2-(2-fluoro-4-iodophenylamino)-6-methoxyphenyl)-1-(2,3-dihydroxypropyl)cyclopropane-1-sulfonamide),PD-0325901(N-[(2R)-2,3-Dihydroxypropoxyl-3,4-difluoro-2-[(2-fluoro-4-iodophenyl)amino]-benzamide),TAK733((R)-3-(2,3-Dihydroxypropyl)-6-fluoro-5-(2-fluoro-4-iodophenylamino)-8-methylpyrido[2,3-d]pyrimidine-4,7(3H,8H)-dione),MEK162/ARRY438162(5-[(4-Bromo-2-fluorophenyl)amino]-4-fluoro-N-(2-hydroxyethoxy)-1-methyl-1H-benzimidazole-6-carboxamide),RO5126766(3-[[3-Fluoro-2-(methylsulfamoylamino)-4-pyridyl]methyl]-4-methyl-7-pyrimidin-2-yloxychromen-2-one),WX-554, RO4987655/CH4987655(3,4-difluoro-2-((2-fluoro-4-iodophenyl)amino)-N-(2-hydroxyethoxy)-5-((3-oxo-1,2-oxazinan-2-yl)methyl)benzamide),or AZD8330(2-((2-fluoro-4-iodophenyl)amino)-N-(2-hydroxyethoxy)-1,5-dimethyl-6-oxo-1,6-dihydropyridine-3-carboxamide).In one embodiment, the cancer or hematological malignancy is DLBCL. Inanother embodiment, the cancer or hematological malignancy is ALL. Inanother embodiment, the cancer or hematological malignancy is CTCL.

In other embodiments, provided herein is a method of treating ormanaging cancer or hematological malignancy comprising administering toa patient a therapeutically effective amount of a compound providedherein (e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37,38, 40, 41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or apharmaceutically acceptable derivative (e.g., salt or solvate) thereof,in combination with an EZH2 inhibitor. In one embodiment, the EZH2inhibitor is EPZ-6438, GSK-126, GSK-343, El1, or 3-deazaneplanocin A(DNNep). In one embodiment, the cancer or hematological malignancy isDLBCL. In another embodiment, the cancer or hematological malignancy isiNHL. In another embodiment, the cancer or hematological malignancy isALL. In another embodiment, the cancer or hematological malignancy isCTCL.

In other embodiments, provided herein is a method of treating ormanaging cancer or hematological malignancy comprising administering toa patient a therapeutically effective amount of a compound providedherein (e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37,38, 40, 41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or apharmaceutically acceptable derivative (e.g., salt or solvate) thereof,in combination with a bcl-2 inhibitor. In one embodiment, the BCL2inhibitor is ABT-199(4-[4-[[2-(4-Chlorophenyl)-4,4-dimethylcyclohex-1-en-1-yl]methyl]piperazin-1-yl]-N-[[3-nitro-4-[[(tetrahydro-2H-pyran-4-yl)methyl]amino]phenyl]sulfonyl]-2-[(1H-pyrrolo[2,3-b]pyridin-5-yl)oxy]benzamide),ABT-737(4-[4-[[2-(4-chlorophenyl)phenyl]methyl]piperazin-1-yl]-N-[4-[[(2R)-4-(dimethylamino)-1-phenylsulfanylbutan-2-yl]amino]-3-nitrophenyl]sulfonylbenzamide),ABT-263((R)-4-(4-44′-chloro-4,4-dimethyl-3,4,5,6-tetrahydro-[1,1′-biphenyl]-2-yl)methyl)piperazin-1-yl)-N-((4-((4-morpholino-1-(phenylthio)butan-2-yl)amino)-3((trifluoromethyl)sulfonyl)phenyl)sulfonyl)benzamide), GX15-070(obatoclax mesylate,(2Z)-2-[(5Z)-5-[(3,5-dimethyl-1H-pyrrol-2-yl)methylidene]-4-methoxypyrrol-2-ylidene]indole;methanesulfonic acid))), or G3139 (Oblimersen). In one embodiment, thecancer or hematological malignancy is DLBCL. In another embodiment, thecancer or hematological malignancy is iNHL. In another embodiment, thecancer or hematological malignancy is CLL. In another embodiment, thecancer or hematological malignancy is ALL. In another embodiment, thecancer or hematological malignancy is CTCL.

In other embodiments, provided herein is a method of treating ormanaging iNHL comprising administering to a patient a therapeuticallyeffective amount of a compound provided herein (e.g., compounds 2, 4, 7,9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73,75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptable derivative(e.g., salt or solvate) thereof, in combination with rituximab. In oneembodiment, the patient is an elderly patient. In another embodiment,iNHL is relapsed or refractory.

In other embodiments, provided herein is a method of treating ormanaging iNHL comprising administering to a patient a therapeuticallyeffective amount of a compound provided herein (e.g., compounds 2, 4, 7,9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73,75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptable derivative(e.g., salt or solvate) thereof, in combination with bendamustine. Inone embodiment, iNHL is relapsed or refractory.

In other embodiments, provided herein is a method of treating ormanaging iNHL comprising administering to a patient a therapeuticallyeffective amount of a compound provided herein (e.g., compounds 2, 4, 7,9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73,75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptable derivative(e.g., salt or solvate) thereof, in combination with rituximab, and infurther combination with bendamustine. In one embodiment, iNHL isrelapsed or refractory.

In other embodiments, provided herein is a method of treating ormanaging iNHL comprising administering to a patient a therapeuticallyeffective amount of a compound provided herein (e.g., compounds 2, 4, 7,9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73,75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptable derivative(e.g., salt or solvate) thereof, in combination with lenalidomide. Inone embodiment, iNHL is relapsed or refractory.

In other embodiments, provided herein is a method of treating ormanaging CLL comprising administering to a patient a therapeuticallyeffective amount of a compound provided herein (e.g., compounds 2, 4, 7,9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73,75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptable derivative(e.g., salt or solvate) thereof, in combination with rituximab. In oneembodiment, the patient is an elderly patient. In another embodiment,CLL is relapsed or refractory.

In other embodiments, provided herein is a method of treating ormanaging CLL comprising administering to a patient a therapeuticallyeffective amount of a compound provided herein (e.g., compounds 2, 4, 7,9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73,75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptable derivative(e.g., salt or solvate) thereof, in combination with bendamustine. Inone embodiment, CLL is relapsed or refractory.

In other embodiments, provided herein is a method of treating ormanaging CLL comprising administering to a patient a therapeuticallyeffective amount of a compound provided herein (e.g., compounds 2, 4, 7,9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73,75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptable derivative(e.g., salt or solvate) thereof, in combination with rituximab, and infurther combination with bendamustine. In one embodiment, CLL isrelapsed or refractory.

In other embodiments, provided herein is a method of treating ormanaging CLL comprising administering to a patient a therapeuticallyeffective amount of a compound provided herein (e.g., compounds 2, 4, 7,9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73,75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptable derivative(e.g., salt or solvate) thereof, in combination with lenalidomide. Inone embodiment, CLL is relapsed or refractory.

In other embodiments, provided herein is a method of treating ormanaging DLBCL comprising administering to a patient a therapeuticallyeffective amount of a compound provided herein (e.g., compounds 2, 4, 7,9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73,75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptable derivative(e.g., salt or solvate) thereof, in combination with rituximab. In oneembodiment, the patient is an elderly patient. In another embodiment,DLBCL is relapsed or refractory.

In other embodiments, provided herein is a method of treating ormanaging DLBCL comprising administering to a patient a therapeuticallyeffective amount of a compound provided herein (e.g., compounds 2, 4, 7,9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73,75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptable derivative(e.g., salt or solvate) thereof, in combination with bendamustine. Inone embodiment, DLBCL is relapsed or refractory.

In other embodiments, provided herein is a method of treating ormanaging DLBCL comprising administering to a patient a therapeuticallyeffective amount of a compound provided herein (e.g., compounds 2, 4, 7,9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73,75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptable derivative(e.g., salt or solvate) thereof, in combination with rituximab, and infurther combination with bendamustine. In one embodiment, DLBCL isrelapsed or refractory.

In other embodiments, provided herein is a method of treating ormanaging DLBCL comprising administering to a patient a therapeuticallyeffective amount of a compound provided herein (e.g., compounds 2, 4, 7,9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73,75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptable derivative(e.g., salt or solvate) thereof, in combination with R-GDP (rituximab,cyclophosphamide, vincristine and prednisone). In one embodiment, DLBCLis relapsed or refractory. In another embodiment, the treatment is donesubsequent to treatment by R-CHOP.

In other embodiments, provided herein is a method of treating ormanaging DLBCL comprising administering to a patient a therapeuticallyeffective amount of a compound provided herein (e.g., compounds 2, 4, 7,9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73,75, 77, 79, 80, 81, and 88), or a pharmaceutically acceptable derivative(e.g., salt or solvate) thereof, in combination with ibrutinib. In oneembodiment, DLBCL is relapsed or refractory.

In other embodiments, provided herein is a method of treating ormanaging T-cell lymphoma (PTCL or CTCL) comprising administering to apatient a therapeutically effective amount of a compound provided herein(e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40,41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or apharmaceutically acceptable derivative (e.g., salt or solvate) thereof,in combination with rituximab. In one embodiment, T-cell lymphoma isrelapsed or refractory.

In other embodiments, provided herein is a method of treating ormanaging T-cell lymphoma (PTCL or CTCL) comprising administering to apatient a therapeutically effective amount of a compound provided herein(e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40,41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or apharmaceutically acceptable derivative (e.g., salt or solvate) thereof,in combination with bendamustine. In one embodiment, T-cell lymphoma isrelapsed or refractory.

In other embodiments, provided herein is a method of treating ormanaging T-cell lymphoma (PTCL or CTCL) comprising administering to apatient a therapeutically effective amount of a compound provided herein(e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40,41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or apharmaceutically acceptable derivative (e.g., salt or solvate) thereof,in combination with rituximab, and in further combination withbendamustine. In one embodiment, T-cell lymphoma is relapsed orrefractory.

In other embodiments, provided herein is a method of treating ormanaging T-cell lymphoma (PTCL or CTCL) comprising administering to apatient a therapeutically effective amount of a compound provided herein(e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40,41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or apharmaceutically acceptable derivative (e.g., salt or solvate) thereof,in combination with romidepsin. In one embodiment, T-cell lymphoma isrelapsed or refractory.

In other embodiments, provided herein is a method of treating ormanaging mantle cell lymphoma comprising administering to a patient atherapeutically effective amount of a compound provided herein (e.g.,compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41,52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or a pharmaceuticallyacceptable derivative (e.g., salt or solvate) thereof, in combinationwith rituximab. In one embodiment, mantle cell lymphoma is relapsed orrefractory.

In other embodiments, provided herein is a method of treating ormanaging mantle cell lymphoma comprising administering to a patient atherapeutically effective amount of a compound provided herein (e.g.,compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41,52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or a pharmaceuticallyacceptable derivative (e.g., salt or solvate) thereof, in combinationwith bendamustine. In one embodiment, mantle cell lymphoma is relapsedor refractory.

In other embodiments, provided herein is a method of treating ormanaging mantle cell lymphoma comprising administering to a patient atherapeutically effective amount of a compound provided herein (e.g.,compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41,52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or a pharmaceuticallyacceptable derivative (e.g., salt or solvate) thereof, in combinationwith rituximab, an din further combination with bendamustine. In oneembodiment, mantle cell lymphoma is relapsed or refractory.

In other embodiments, provided herein is a method of treating ormanaging mantle cell lymphoma comprising administering to a patient atherapeutically effective amount of a compound provided herein (e.g.,compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41,52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or a pharmaceuticallyacceptable derivative (e.g., salt or solvate) thereof, in combinationwith ibrutinib. In one embodiment, mantle cell lymphoma is relapsed orrefractory.

Further, without being limited by a particular theory, it was found thatcancer cells exhibit differential sensitivity profiles to doxorubicinand compounds provided herein. Thus, provided herein is a method oftreating or managing cancer or hematological malignancy comprisingadministering to a patient a therapeutically effective amount of acompound provided herein (e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26,27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81,and 88), or a pharmaceutically acceptable derivative (e.g., salt orsolvate) thereof, in combination with a doxorubicin. In one embodiment,the cancer or hematological malignancy is ALL.

In some embodiments, provided herein is a method of treating or managingcancer or hematological malignancy comprising administering to a patienta therapeutically effective amount of a compound provided herein (e.g.,compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41,52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or a pharmaceuticallyacceptable derivative (e.g., salt or solvate) thereof, in combinationwith a AraC. In one embodiment, the cancer or hematological malignancyis AML.

In specific embodiments, compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30,32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88or a pharmaceutically acceptable form thereof, is used in combinationwith one or more second agent or second therapy provided herein.

In some embodiments, the second agent is an antibody-drug conjugate,such as, e.g., inotuzumab ozogamicin, or brentuximab vedotin.

In some embodiments, the second agent is a cytotoxic agent, such as,e.g., bendamustine, gemcitabine, oxaliplatin, cyclophosphamide,vincristine, vinblastine, anthracycline (e.g., daunorubicin ordaunomycin, doxorubicin), actinomycin, dactinomycin, bleomycin,clofarabine, nelarabine, cladribine, asparaginase, methotrexate, orpralatrexate.

In some embodiments, the second agent is one or more other anti-canceragents or chemotherapeutic agents, such as, e.g., fludarabine,ibrutinib, fostamatinib, lenalidomide, thalidomide, rituximab,cyclophosphamide, doxorubicin, vincristine, prednisone, or R-CHOP(Rituximab, Cyclophosphamide, Doxorubicin or Hydroxydaunomycin,Vincristine or Oncovin, Prednisone).

In some embodiments, the second agent is an antibody for a cytokine(e.g., an IL-15 antibody, an IL-21 antibody, an IL-4 antibody, an IL-7antibody, an IL-2 antibody, an IL-9 antibody). In some embodiments, thesecond agent is a JAK1 inhibitor, a JAK3 inhibitor, a pan-JAK inhibitor,a BTK inhibitor, an SYK inhibitor, or a PI3K delta inhibitor. In someembodiments, the second agent is an antibody for a chemokine.

Without being limited to a particular theory, a targeted combinationtherapy described herein has reduced side effect and/or enhancedefficacy. For example, in one embodiment, provided herein is acombination therapy for treating CLL with a compound described herein(e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40,41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88) and a second activeagent (e.g., IL-15 antibodies, IL-21 antibodies, IL-4 antibodies, IL-7antibodies, IL-2 antibodies, IL-9 antibodies, JAK1 inhibitors, JAK3inhibitors, pan-JAK inhibitors, BTK inhibitors, SYK inhibitors, and/orPI3K delta inhibitors).

Further without being limited by a particular theory, it was found thata compound provided herein (e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26,27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81,and 88) does not affect BTK or MEK pathway. Accordingly, in someembodiments, provided herein is a method of treating or managing canceror hematological malignancy comprising administering to a patient atherapeutically effective amount of a compound provided herein (e.g.,compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41,52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or a pharmaceuticallyacceptable derivative (e.g., salt or solvate) thereof, in combinationwith a BTK inhibitor. In one embodiment, the BTK inhibitor is ibrutinib.In one embodiment, the BTK inhibitor is AVL-292. In one embodiment, thecancer or hematological malignancy is DLBCL. In another embodiment, thecancer or hematological malignancy is CLL.

In other embodiments, provided herein is a method of treating ormanaging cancer or hematological malignancy comprising administering toa patient a therapeutically effective amount of a compound providedherein (e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37,38, 40, 41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or apharmaceutically acceptable derivative (e.g., salt or solvate) thereof,in combination with a MEK inhibitor. In one embodiment, the MEKinhibitor is tametinib, selumetinob, AS703026/MSC1935369,XL-518/GDC-0973, BAY869766/RDEA119, GSK1120212 (trametinib), pimasertib,refametinib, PD-0325901, TAK733, MEK162/ARRY438162, R05126766, WX-554,RO4987655/CH4987655 or AZD8330. In one embodiment, the cancer orhematological malignancy is DLBCL. In another embodiment, the cancer orhematological malignancy is ALL. In another embodiment, the cancer orhematological malignancy is CTCL.

In other embodiments, provided herein is a method of treating ormanaging cancer or hematological malignancy comprising administering toa patient a therapeutically effective amount of a compound providedherein (e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37,38, 40, 41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or apharmaceutically acceptable derivative (e.g., salt or solvate) thereof,in combination with a bcl-2 inhibitor. In one embodiment, the BCL2inhibitor is ABT-199, ABT-737, ABT-263, GX15-070 (obatoclax mesylate) orG3139 (Genasense). In one embodiment, the cancer or hematologicalmalignancy is DLBCL. In another embodiment, the cancer or hematologicalmalignancy is ALL. In another embodiment, the cancer or hematologicalmalignancy is CTCL.

Further, without being limited by a particular theory, it was found thatcancer cells exhibit differential sensitivity profiles to doxorubicinand compounds provided herein. Thus, provided herein is a method oftreating or managing cancer or hematological malignancy comprisingadministering to a patient a therapeutically effective amount of acompound provided herein (e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26,27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81,and 88), or a pharmaceutically acceptable derivative (e.g., salt orsolvate) thereof, in combination with a doxorubicin. In one embodiment,the cancer or hematological malignancy is ALL.

In some embodiments, provided herein is a method of treating or managingcancer or hematological malignancy comprising administering to a patienta therapeutically effective amount of a compound provided herein (e.g.,compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41,52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88), or a pharmaceuticallyacceptable derivative (e.g., salt or solvate) thereof, in combinationwith a AraC. In one embodiment, the cancer or hematological malignancyis AML.

In specific embodiments, compounds 2, 4, 7, 9, 17, 19, 21, 26, 27, 30,32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81, and 88or a pharmaceutically acceptable form thereof, is used in combinationwith one or more second agent or second therapy provided herein.

In certain embodiments, provided herein are pharmaceutical compositioncomprising a therapeutically effective amount of a compound providedherein (e.g., a compound of any of Formulae (I″), (I′), (A′), (I), (A),(II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII),(XIV), (XV), (XVI), and (XVII), e.g., a selective PI3K-γ inhibitor,e.g., Compound 4), or a pharmaceutically acceptable form thereof, and aPI3K-delta inhibitor.

In one embodiment, the PI3K-delta inhibitor is a PI3K-delta selectiveinhibitor. In one embodiment, the PI3K-delta inhibitor is GS-1101(Cal-101), GSK-2269557, GS-9820, AMG319, or TGR-1202, or a mixturethereof. In one embodiment, the PI3K-delta inhibitor is of the formula:

or a pharmaceutically acceptable form thereof.

In one embodiment, the molar ratio of the compound, or apharmaceutically acceptable form thereof, to the PI3K-delta inhibitor isin the range of from about 10000:1 to about 1:10000. In one embodiment,the molar ratio of the compound, or a pharmaceutically acceptable formthereof, to the PI3K-delta inhibitor is in the range of from about 10:1to about 1:10. In one embodiment, the composition comprises thecompound, or a pharmaceutically acceptable form thereof, at an amount ofin the range of from about 0.01 mg to about 75 mg and the PI3K-deltainhibitor at an amount of in the range of from about 0.01 mg to about1100 mg. In one embodiment, the compound, or a pharmaceuticallyacceptable form thereof, and the PI3K-delta inhibitor are the onlytherapeutically active ingredients.

In one embodiment, the compound, or pharmaceutically acceptable formthereof, and the PI3K-delta inhibitor are in a single dosage form. Inone embodiment, the compound, or pharmaceutically acceptable formthereof, and the PI3K-delta inhibitor are in separate dosage forms. Inone embodiment, the composition further comprising a pharmaceuticallyacceptable excipient.

In one embodiment, the composition is synergistic in treating a cancer,inflammatory disease, or autoimmune disease.

In one embodiment, provided herein is a method of treating aPI3K-mediated disorder in a subject, comprising administering to thesubject a therapeutically effective amount of the composition.

In certain embodiments, provided herein are methods treating aPI3K-mediated disorder in a subject, comprising administering to thesubject a therapeutically effective amount of a compound provided herein(e.g., a compound of any of Formulae (I″), (I′), (A′), (I), (A), (II),(III), (IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII), (XIV),(XV), (XVI), and (XVII), e.g., a selective PI3K-γ inhibitor, e.g.,Compound 4), or a pharmaceutically acceptable form thereof, incombination with a PI3K-delta inhibitor.

In one embodiment, the compound, or a pharmaceutically acceptable formthereof, is administered concurrently with the PI3K-delta inhibitor. Inone embodiment, the compound, or a pharmaceutically acceptable formthereof, is administered subsequent to the PI3K-delta inhibitor. In oneembodiment, the compound, or a pharmaceutically acceptable form thereof,is administered prior to the PI3K-delta inhibitor. In one embodiment,the compound, or a pharmaceutically acceptable form thereof, isadministered alone after discontinuing the administration of thePI3K-delta inhibitor.

In one embodiment, the PI3K-mediated disorder is a cancer, autoimmunedisease, or inflammatory disease. In one embodiment, the cancer is ofhematopoietic origin. In one embodiment, the cancer is a leukemia orlymphoma. In one embodiment, the leukemia or lymphoma is a B-celllymphoma, T-cell lymphoma, non-Hodgkin lymphoma, Hodgkin lymphoma, oranaplastic large cell lymphoma.

In one embodiment, the cancer is a solid tumor. In one embodiment, thecancer is selected from one or more of: a cancer of the pulmonarysystem, a brain cancer, a cancer of the gastrointestinal tract, a skincancer, a genitourinary cancer, a pancreatic cancer, a lung cancer, amedullobastoma, a basal cell carcinoma, a glioma, a breast cancer, aprostate cancer, a testicular cancer, an esophageal cancer, ahepatocellular cancer, a gastric cancer, a gastrointestinal stromaltumor (GIST), a colon cancer, a colorectal cancer, an ovarian cancer, amelanoma, a neuroectodermal tumor, head and neck cancer, a sarcoma, asoft-tissue sarcoma, fibrosarcoma, myxosarcoma, liposarcoma, achondrosarcoma, an osteogenic sarcoma, a chordoma, an angiosarcoma, anendotheliosarcoma, a lymphangiosarcoma, a lymphangioendotheliosarcoma, asynovioma, a mesothelioma, a leiomyosarcoma, a cervical cancer, auterine cancer, an endometrial cancer, a carcinoma, a bladder carcinoma,an epithelial carcinoma, a squamous cell carcinoma, an adenocarcinoma, abronchogenic carcinoma, a renal cell carcinoma, a hepatoma, a bile ductcarcinoma, a neuroendocrine cancer, a carcinoid tumor, diffuse typegiant cell tumor, and glioblastoma.

In one embodiment, the PI3K-delta inhibitor is a PI3K-delta selectiveinhibitor. In one embodiment, the PI3K-delta inhibitor is of theformula:

or a pharmaceutically acceptable form thereof.

In one embodiment, the compound, or a pharmaceutically acceptable formthereof, and the PI3K-delta inhibitor are in a single dosage form. Inone embodiment, the compound, or a pharmaceutically acceptable formthereof, and the PI3K-delta inhibitor are in separate dosage forms.

In one embodiment, the concentration of the compound that is required toachieve 50% inhibition is at least 20%, 30%, 40%, or 50% lower when thecompound is administered in combination with the PI3K-delta inhibitorthan when the compound is administered individually. In one embodiment,the concentration of the PI3K-delta inhibitor that is required toachieve 50% inhibition is at least 20%, 30%, 40%, or 50% lower when thePI3K-delta inhibitor is administered in combination with the compoundthan when the PI3K-delta inhibitor is administered individually. In oneembodiment, the dose of the compound that is required to achieve 50%inhibition is at least 20%, 30%, 40%, or 50% lower when the compound isadministered in combination with the PI3K-delta inhibitor than when thecompound is administered individually. In one embodiment, the dose ofthe PI3K-delta inhibitor that is required to achieve 50% inhibition isat least 20%, 30%, 40%, or 50% lower when the PI3K-delta inhibitor isadministered in combination with the compound than when the PI3K-deltainhibitor is administered individually.

In one embodiment, the combination is synergistic as indicated by acombination index value that is less than 0.7, 0.5, or 0.1 for thecombination of the compound and the PI3K-delta inhibitor. In oneembodiment, the combination index value is assessed at 50% inhibition.In one embodiment, the combination index value is assessed at 50% growthinhibition. In one embodiment, the combination is synergistic asindicated by a Synergy Score that is greater than 1, 2, or 3 for thecombination of the Compound 4 and the PI3K-delta inhibitor. In oneembodiment, the combination is synergistic as indicated by a SynergyScore that is greater than 1, 2, or 3, for the combination of thecompound and the PI3K-delta inhibitor for inhibition or growthinhibition.

In one embodiment, the PI3K-mediated disorder is cancer, and theanti-cancer effect provided by the combination is at least 2 foldgreater, at least 3 fold greater, at least 5 fold greater, or at least10 fold greater than the anti-cancer effect provided by Compound 4, orpharmaceutically acceptable form thereof, alone. In one embodiment, thePI3K-mediated disorder is cancer, and the anti-cancer effect provided bythe combination is at least 2 fold greater, at least 3 fold greater, atleast 5 fold greater, or at least 10 fold greater than the anti-cancereffect provided by the PI3K-delta inhibitor alone.

In one embodiment, wherein one or more side effects associated withadministration of the compound, or a pharmaceutically acceptable formthereof, alone is reduced when the combination is administered at a dosethat achieves the same therapeutic effect. In one embodiment, one ormore side effects associated with administration of the PI3K-deltainhibitor alone is reduced when the combination is administered at adose that achieves the same therapeutic effect.

Combinations with Immune Modulators

While not wishing to be bound by theory, it is believed that tumorgrowth is influenced by at least two classes of immune cells in thetumor microenvironment: effector cells (including cytotoxic cells and M1macrophages) which have anti-tumor activity, and tumor associatedsuppressor cells (including M2 macrophages, MDSC, Tregs, and regulatorydendritic cells) which have pro-tumor activity because they inhibit theeffector cells or provide direct growth stimulation to the tumor cellsor tumor vasculature. An abundance of suppressor cells can lead to tumorimmune tolerance, and enhancement of tumor growth. A combination cancertherapy can be designed taking this mechanism into consideration.

For example, in embodiments, a PI3K-γ inhibitor as described herein (ora compound provided herein (e.g., compounds 2, 4, 7, 9, 17, 19, 21, 26,27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73, 75, 77, 79, 80, 81,and 88)) is administered in combination with a second therapeutic thatblocks homeostatic down-regulation of a T cell response in an effector Tcell. This second agent may be an immune checkpoint therapy as describedbelow. As another example, in embodiments, a PI3K-γ inhibitor asdescribed herein is administered in combination with a secondtherapeutic that reduces or eliminates suppressive cells in the tumormicroenvironment, e.g., may deplete MDSCs, TAMs or M2 macrophages, orany combination thereof. This agent could comprise, e.g., a CSF1Rinhibitor, a CCL2 inhibitor, a CXCR4 inhibitor, a MEK inhibitor, or anMTOR inhibitor, or any combination thereof. In some embodiments, thesecond agent is an immunotherapy such as a tumor vaccine, e.g., a tumorvaccine described herein. In some embodiments, the second agent is acell therapy, e.g., a dendritic cell or a chimeric T cell, e.g., asdescribed herein. In some embodiments, the second agent is aninterleukin, e.g., IL7, IL12, IL15, or IL21. According to non-limitingtheory, some interleukins exert an anti-cancer effect by stimulating thegrowth of immune cell populations.

In another embodiment, a compound provided herein (e.g., compounds 2, 4,7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63,73, 75, 77, 79, 80, 81, and 88) is administered in combination with avaccine, e.g., a cancer vaccine, (e.g., a dendritic cell renal carcinoma(DC-RCC) vaccine). In certain embodiments, the combination of compoundand the DC-RCC vaccine is used to treat a cancer, e.g., a cancer asdescribed herein (e.g., a renal carcinoma, e.g., metastatic renal cellcarcinoma (RCC) or clear cell renal cell carcinoma (CCRCC)).

In some embodiments, a compound provided herein, or a pharmaceuticallyacceptable derivative (e.g., salt or solvate) thereof, is administeredin combination with one or more immune checkpoint therapies. In someembodiments, provided herein is a method of treating a cancer in asubject, comprising administering to the subject a PI3K gamma inhibitoror a compound as described herein (e.g., a compound of any of Formulae(I″), (I′), (A′), (I), (A), (II), (III), (IV), (V), (VI), (VII), (IX),(X), (XI), (XII), (XIII), (XIV), (XV), (XVI), and (XVII), e.g., Compound4, as described herein) in combination with one or more immunecheckpoint therapies (e.g., PD-1 or PD-L1 inhibitors). In someembodiments, provided herein is a method of treating a solid cancer in asubject, comprising administering to the subject Compound 4, or apharmaceutically acceptable form thereof, in combination with one ormore of PD-1 or PD-L1 inhibitors. In one embodiment, the cancer ismelanoma, bladder cancer, head and neck cancer, lung cancer (e.g.,non-small cell lung cancer), or renal cell carcinoma. In one embodiment,the cancer is melanoma. In one embodiment, the cancer is bladder cancer.In one embodiment, the cancer is lung cancer. In one embodiment, thecancer is non-small cell lung cancer. In one embodiment, the cancer isrenal cell carcinamo. In one embodiment, the cancer is head and neckcancer. In one embodiment, the cancer is breast cancer. In oneembodiment, the cancer is triple-negative breast cancer. In oneembodiment, the cancer is colon cancer. In one embodiment, the cancer isglioblastoma. In one embodiment, the cancer is ovarian cancer.

In some embodiments, the subject is naive to immunotherapy treatment. Insome embodiments, the subject is naive to radiation therapy treatment.In some embodiments, the subject is naive to chemotherapy treatment.

In some embodiments, the subject has been pre-treated or previouslytreated with one or more immunotherapy treatments. In one embodiment,the subject is responsive to the pre-treatment or previous treatmentwith the immunotherapy. In one embodiment, the immunotherapy treatmentis a checkpoint treatment such as a PD-1 or PD-L1 inhibitor. In oneembodiment, the subject is a smoker.

In one embodiment, the cancer is melanoma, and the subject has beenpre-treated or previously treated with one or more immunotherapytreatments. In one embodiment, the subject has been pre-treated orpreviously treated with two or more immunotherapy treatments.

In one embodiment, the cancer is head and neck cancer, lung cancer(e.g., non-small cell lung cancer), renal cell carcinoma, or bladdercancer, and the subject has been pre-treated or previously treated withone immunotherapy treatment.

In one embodiment, the cancer is breast cancer (e.g., triple-negativebreast cancer), ovarian cancer, glioblastoma, or colon cancer, and thesubject is naive to immunotherapy treatment.

In some embodiments, the immune checkpoint therapy inhibits CTLA-4,PD-1, or PD-L1, or any combination thereof. The immune checkpointtherapy may be, e.g., a small molecule or an antibody. In someembodiments, the immune checkpoint therapy is an antibody that inhibitsprogrammed cell death 1 (also known as PD-1). In another embodiment, theimmune checkpoint therapy is nivolumab (also known as Opdivo). In someembodiments, the immune checkpoint therapy is anti-PD-L1 (programmedcell death ligand 1, also known as cluster of differentiation 274(CD274)), anti-PDL2, or anti-CTLA-4 (cytotoxic T-lymphocyte antigen 4,also known as cluster of differentiation (CD152)) antibody. Certainanti-PD-1, anti-PD-L1, and anti-CTLA-4 antibodies have activity inpreclinical and clinical tumor models. Cancer Res; 73(12) Jun. 15, 2013;Curran M A et al. PNAS 2010; 107:4275-4280; Topalian et al. N Engl J Med2012; 366:2443-2454; Wolchok et al., 2013. NEJM 369.

There are two main types of immune checkpoint therapies: an activator ofa costimulatory molecule, and an inhibitor of an immune checkpointmolecule.

When the immune checkpoint therapy is an activator of a costimulatorymolecule, it may be, e.g., chosen from an agonist (e.g., an agonisticantibody or antigen-binding fragment thereof, or a soluble fusion) ofOX40, CD2, CD27, CDS, ICAM-1, LFA-1 (CD11a/CD18), ICOS (CD278), 4-1BB(CD137), GITR, CD30, CD40, BAFFR, HVEM, CD7, LIGHT, NKG2C, SLAMF7,NKp80, CD160, B7-H3 or CD83 ligand. In certain embodiments, the immunecheckpoint therapy is an inhibitor of OX40 or anti-OX40 ab.

In the second situation, the immune checkpoint therapy is an inhibitorof an immune checkpoint molecule, for instance, an inhibitor of PD-1,PD-L1, PD-L2, CTLA4, TIM3, LAG3, VISTA, BTLA, TIGIT, LAIR1, CD160, 2B4and/or TGFR beta. For instance, the inhibitor of an immune checkpointmolecule may inhibit PD-1, PD-L1, LAG-3, TIM-3 or CTLA4, or anycombination thereof.

Inhibition of an inhibitory molecule can be performed at the DNA, RNA orprotein level. For example, an inhibitory nucleic acid (e.g., a dsRNA,siRNA or shRNA), can be used to inhibit expression of an inhibitorymolecule. In other embodiments, the inhibitor of an inhibitory signalis, a polypeptide e.g., a soluble ligand (e.g., PD-1-Ig or CTLA-4 Ig),or an antibody or antigen-binding fragment thereof, that binds to theinhibitory molecule; e.g., an antibody or fragment thereof (alsoreferred to herein as “an antibody molecule”) that binds to PD-1, PD-L1,PD-L2, CTLA4, TIM3, LAG3, VISTA, BTLA, TIGIT, LAIR1, CD160, 2B4 and/orTGFR beta, or a combination thereof.

The antibody molecule may be, e.g., a full antibody or fragment thereof(e.g., a Fab, F(ab′)₂, Fv, or a single chain Fv fragment (scFv)). Theantibody molecule may be, e.g., in the form of a bispecific antibodymolecule. In one embodiment, the bispecific antibody molecule has afirst binding specificity to PD-1 or PD-L1 and a second bindingspecifity, e.g., a second binding specificity to TIM-3, LAG-3, or PD-L2.In certain embodiments, the antibody molecule is administered byinjection (e.g., subcutaneously or intravenously) at a dose of about 1to 30 mg/kg, e.g., about 5 to 25 mg/kg, about 10 to 20 mg/kg, about 1 to5 mg/kg, or about 3 mg/kg. The dosing schedule can vary from e.g., oncea week to once every 2, 3, or 4 weeks.

In certain embodiments, the immune checkpoint therapy is an inhibitor ofPD-1, e.g., human PD-1. In another embodiment, the immune checkpointtherapy is an inhibitor of PD-L1, e.g., human PD-L1. In one embodiment,the inhibitor of PD-1 or PD-L1 is an antibody molecule to PD-1 or PD-L1.The PD-1 or PD-L1 inhibitor can be administered alone, or in combinationwith other immune checkpoint therapies, e.g., in combination with aninhibitor of LAG-3, TIM-3 or CTLA4. In some embodiments, the inhibitorof PD-1 or PD-L1, e.g., the anti-PD-1 or PD-L1 antibody molecule, isadministered in combination with a LAG-3 inhibitor, e.g., an anti-LAG-3antibody molecule. In another embodiment, the inhibitor of PD-1 orPD-L1, e.g., the anti-PD-1 or PD-L1 antibody molecule, is administeredin combination with a TIM-3 inhibitor, e.g., an anti-TIM-3 antibodymolecule. In yet other embodiments, the inhibitor of PD-1 or PD-L1,e.g., the anti-PD-1 antibody molecule, is administered in combinationwith a LAG-3 inhibitor, e.g., an anti-LAG-3 antibody molecule, and aTIM-3 inhibitor, e.g., an anti-TIM-3 antibody molecule. Othercombinations of immune checkpoint therapies with a PD-1 inhibitor (e.g.,one or more of PD-L2, CTLA4, TIM3, LAG3, VISTA, BTLA, TIGIT, LAIR1,CD160, 2B4 and/or TGFR) are also within the present invention. Any ofthe PI3K inhibitor molecules known in the art or disclosed herein can beused in the aforesaid combinations of inhibitors of checkpoint molecule.

In some embodiments, the anti-PD-1 antibody is Nivolumab. Alternativenames for Nivolumab include MDX-1106, MDX-1106-04, ONO-4538, orBMS-936558. In some embodiments, the anti-PD-1 antibody is Nivolumab(CAS Registry Number: 946414-94-4). Nivolumab is a fully human IgG4monoclonal antibody which specifically blocks PD1. Nivolumab (clone 5C4)and other human monoclonal antibodies that specifically bind to PD1 aredisclosed in U.S. Pat. No. 8,008,449 and WO2006/121168.

In other embodiments, the anti-PD-1 antibody is Pembrolizumab.Pembrolizumab (Trade name KEYTRUDA formerly Lambrolizumab, also known asMerck 3745, MK-3475 or SCH-900475) is a humanized IgG4 monoclonalantibody that binds to PD1. Pembrolizumab is disclosed, e.g., in Hamid,O. et al. (2013) New England Journal of Medicine 369 (2): 134-44,WO2009/114335, and U.S. Pat. No. 8,354,509.

In some embodiments, the anti-PD-1 antibody is Pidilizumab. Pidilizumab(CT-011; Cure Tech) is a humanized IgG1k monoclonal antibody that bindsto PD1. Pidilizumab and other humanized anti-PD-1 monoclonal antibodiesare disclosed in WO2009/101611. Other anti-PD1 antibodies are disclosedin U.S. Pat. No. 8,609,089, US 2010028330, and/or US 20120114649. Otheranti-PD1 antibodies include AMP 514 (Amplimmune).

In some embodiments, the PD-1 inhibitor is an immunoadhesin (e.g., animmunoadhesin comprising an extracellular or PD-1 binding portion ofPD-L1 or PD-L2 fused to a constant region (e.g., an Fc region of animmunoglobulin sequence)). In some embodiments, the PD-1 inhibitor isAMP-224. In some embodiments, a PI3K inhibitor, e.g., a PI3K-γ inhibitoras described herein (e.g., Compound 4), is administered together with animmunoadhesin (e.g., an immunoadhesin comprising an extracellular orPD-1 binding portion of PD-L1 or PD-L2 fused to a constant region (e.g.,an Fc region of an immunoglobulin sequence)). In some embodiments, thecombination therapy is used in a method of treating a cancer, asdescribed herein.

In some embodiments, the PD-L1 inhibitor is anti-PD-L1 antibody. In someembodiments, the anti-PD-L1 inhibitor is chosen from YW243.55.570,MPDL3280A, MEDI-4736, MSB-0010718C, or MDX-1105.

In one embodiment, the PD-L1 inhibitor is MDX-1105. MDX-1105, also knownas BMS-936559, is an anti-PD-L1 antibody described in WO2007/005874.

In one embodiment, the PD-L1 inhibitor is YW243.55.570. The YW243.55.570antibody is an anti-PD-L1 described in WO 2010/077634 (heavy and lightchain variable region sequences shown in SEQ ID Nos. 20 and 21,respectively).

In one embodiment, the PD-L1 inhibitor is MDPL3280A (Genentech/Roche).MDPL3280A is a human Fc optimized IgG1 monoclonal antibody that binds toPD-L1. MDPL3280A and other human monoclonal antibodies to PD-L1 aredisclosed in U.S. Pat. No. 7,943,743 and U.S Publication No.:20120039906.

In other embodiments, the PD-L2 inhibitor is AMP-224. AMP-224 is a PD-L2Fc fusion soluble receptor that blocks the interaction between PD1 andB7-H1 (B7-DCIg; Amplimmune; e.g., disclosed in WO2010/027827 andWO2011/066342).

In one embodiment, the LAG-3 inhibitor is an anti-LAG-3 antibodymolecule. In one embodiment, the LAG-3 inhibitor is BMS-986016.

In some embodiments, the anti-PD-L1 binding antagonist is chosen fromYW243.55.570, MPDL3280A, MEDI-4736, MSB-0010718C, or MDX-1105. MDX-1105,also known as BMS-936559, is an anti-PD-L1 antibody described inWO2007/005874. Antibody YW243.55.570 (heavy and light chain variableregion sequences shown in SEQ ID Nos. 20 and 21, respectively) is ananti-PD-L1 described in WO 2010/077634.

In some embodiments, the anti-PD-1 antibody is Nivolumab. Alternativenames for Nivolumab include MDX-1106, MDX-1106-04, ONO-4538, orBMS-936558. In some embodiments, the anti-PD-1 antibody is Nivolumab(CAS Registry Number: 946414-94-4). Nivolumab (also referred to asBMS-936558 or MDX1106; Bristol-Myers Squibb) is a fully human IgG4monoclonal antibody which specifically blocks PD-1. Nivolumab (clone5C4) and other human monoclonal antibodies that specifically bind toPD-1 are disclosed in U.S. Pat. No. 8,008,449, EP2161336 andWO2006/121168.

In some embodiments, the anti-PD-1 antibody is Pembrolizumab.Pembrolizumab (also referred to as Lambrolizumab, MK-3475, MK03475,SCH-900475 or KEYTRUDA®; Merck) is a humanized IgG4 monoclonal antibodythat binds to PD-1. Pembrolizumab and other humanized anti-PD-1antibodies are disclosed in Hamid, O. et al. (2013) New England Journalof Medicine 369 (2): 134-44, U.S. Pat. No. 8,354,509 and WO2009/114335.

Pidilizumab (CT-011; Cure Tech) is a humanized IgG1k monoclonal antibodythat binds to PD1. Pidilizumab and other humanized anti-PD-1 monoclonalantibodies are disclosed in WO2009/101611.

Other anti-PD1 antibodies include AMP 514 (Amplimmune), among others,e.g., anti-PD1 antibodies disclosed in U.S. Pat. No. 8,609,089, US2010028330, and/or US 20120114649.

In some embodiments, the anti-PD-L1 antibody is MSB0010718C. MSB0010718C(also referred to as A09-246-2; Merck Serono) is a monoclonal antibodythat binds to PD-L1. Pembrolizumab and other humanized anti-PD-L1antibodies are disclosed in WO2013/079174.

MDPL3280A (Genentech/Roche) is a human Fc optimized IgG1 monoclonalantibody that binds to PD-L1. MDPL3280A and other human monoclonalantibodies to PD-L1 are disclosed in U.S. Pat. No. 7,943,743 and U.SPublication No.: 20120039906. Other anti-PD-L1 binding agents includeYW243.55.570 (heavy and light chain variable regions are shown in SEQ IDNOs 20 and 21 in WO2010/077634) and MDX-1105 (also referred to asBMS-936559, and, e.g., anti-PD-L1 binding agents disclosed inWO2007/005874).

AMP-224 (B7-DCIg; Amplimmune; e.g., disclosed in WO2010/027827 andWO2011/066342), is a PD-L2 Fc fusion soluble receptor that blocks theinteraction between PD1 and B7-H1.

In some embodiments, the anti-LAG-3 antibody is BMS-986016. BMS-986016(also referred to as BMS986016; Bristol-Myers Squibb) is a monoclonalantibody that binds to LAG-3. BMS-986016 and other humanized anti-LAG-3antibodies are disclosed in US 2011/0150892, WO2010/019570, andWO2014/008218.

In certain embodiments, the combination therapies disclosed hereininclude a modulator of a costimulatory molecule or an inhibitorymolecule, e.g., a co-inhibitory ligand or receptor.

In one embodiment, the costimulatory modulator, e.g., agonist, of acostimulatory molecule is chosen from an agonist (e.g., an agonisticantibody or antigen-binding fragment thereof, or soluble fusion) ofOX40, CD2, CD27, CDS, ICAM-1, LFA-1 (CD11a/CD18), ICOS (CD278), 4-1BB(CD137), GITR, CD30, CD40, BAFFR, HVEM, CD7, LIGHT, NKG2C, SLAMF7,NKp80, CD160, B7-H3 or CD83 ligand.

In another embodiment, the combination therapies disclosed hereininclude a costimulatory molecule, e.g., an agonist associated with apositive signal that includes a costimulatory domain of CD28, CD27, ICOSand GITR.

Exemplary GITR agonists include, e.g., GITR fusion proteins andanti-GITR antibodies (e.g., bivalent anti-GITR antibodies), such as, aGITR fusion protein described in U.S. Pat. No. 6,111,090, EuropeanPatent No.: 090505B1, U.S. Pat. No. 8,586,023, PCT Publication Nos.: WO2010/003118 and 2011/090754, or an anti-GITR antibody described, e.g.,in U.S. Pat. No. 7,025,962, European Patent No.: 1947183B1, U.S. Pat.Nos. 7,812,135, 8,388,967, 8,591,886, European Patent No.: EP 1866339,PCT Publication No.: WO 2011/028683, PCT Publication No.:WO 2013/039954,PCT Publication No.: WO2005/007190, PCT Publication No.: WO 2007/133822,PCT Publication No.: WO2005/055808, PCT Publication No.: WO 99/40196,PCT Publication No.: WO 2001/03720, PCT Publication No.: WO99/20758, PCTPublication No.: WO2006/083289, PCT Publication No.: WO 2005/115451,U.S. Pat. No. 7,618,632, and PCT Publication No.: WO 2011/051726.

In one embodiment, the inhibitor is a soluble ligand (e.g., aCTLA-4-Ig), or an antibody or antibody fragment that binds to PD-L1,PD-L2 or CTLA4. For example, a compound disclosed herein, e.g., Compound4, can be administered in combination with an anti-CTLA-4 antibody,e.g., ipilimumab, for example, to treat a cancer (e.g., a cancer chosenfrom: a melanoma, e.g., a metastatic melanoma; a lung cancer, e.g., anon-small cell lung carcinoma; or a prostate cancer). Exemplaryanti-CTLA4 antibodies include Tremelimumab (IgG2 monoclonal antibodyavailable from Pfizer, formerly known as ticilimumab, CP-675,206); andIpilimumab (CTLA-4 antibody, also known as MDX-010, Yervoy, CAS No.477202-00-9). In some embodiments, a compound provided herein isadministered in combination with an anti-PD-L1 inhibitor (e.g.,nivolumab) and a CTLA-4 antibody (e.g., ipilimumab). In someembodiments, a compound provided herein is administered in combinationwith nivolumab and ipilimumab.

In some embodiments, a compound provided herein, or a pharmaceuticallyacceptable derivative (e.g., salt or solvate) thereof, is administeredin combination with an anti-PD-L1 or anti-CTLA-4 antibody. In someembodiments, a compound provided herein, or a pharmaceuticallyacceptable derivative (e.g., salt or solvate) thereof, is administeredin combination with an anti-PD-L1 antibody. In another embodiment, acompound provided herein, or a pharmaceutically acceptable derivative(e.g., salt or solvate) thereof, is administered in combination withanti-CTLA-4 antibody. In some embodiments, the anti-PD-L1 antibody isselected from BMS-936559, MPDL3280A, and MDX-1105. In some embodiments,the anti-CTLA-4 antibody is selected from ipilimumab and tremelimumab.

In some embodiments, provided herein is a method of treating breastcancer, colon cancer, pancreatic cancer, melanoma, glioblastoma, or lungcancer comprising administering to a patient a therapeutically effectiveamount of a compound provided herein (e.g., compounds 2, 4, 7, 9, 17,19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73, 75, 77,79, 80, 81, and 88), or a pharmaceutically acceptable derivative (e.g.,salt or solvate) thereof, in combination with an anti-PD-L1 or ananti-CTLA-4 antibody. In another embodiment, the cancer is chosen form acarcinoma (e.g., advanced or metastatic carcinoma), melanoma or a lungcarcinoma, e.g., a non-small cell lung carcinoma. In one embodiment, thecancer is a lung cancer, e.g., a non-small cell lung cancer. In oneembodiment, the cancer is a melanoma, e.g., an advanced melanoma. In oneembodiment, the cancer is an advanced or unresectable melanoma that doesnot respond to other therapies. In other embodiments, the cancer is amelanoma with a BRAF mutation (e.g., a BRAF V600E mutation). In anotherembodiment, the cancer is a hepatocarcinoma, e.g., an advancedhepatocarcinoma, with or without a viral infection, e.g., a chronicviral hepatitis. In another embodiment, the cancer is a prostate cancer,e.g., an advanced prostate cancer. In yet another embodiment, the canceris a myeloma, e.g., multiple myeloma. In yet another embodiment, thecancer is a renal cancer, e.g., a renal cell carcinoma (RCC) (e.g., ametastatic RCC or clear cell renal cell carcinoma (CCRCC)).

For example, a compound provided herein (e.g., compounds 2, 4, 7, 9, 17,19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61, 63, 73, 75, 77,79, 80, 81, and 88) can be administered in combination with ananti-CTLA-4 antibody, e.g., ipilimumab, for example, to treat a cancer(e.g., a cancer chosen from: a melanoma, e.g., a metastatic melanoma; alung cancer, e.g., a non-small cell lung carcinoma; or a prostatecancer). In one embodiment, a compound provided herein (e.g., compounds2, 4, 7, 9, 17, 19, 21, 26, 27, 30, 32, 35, 37, 38, 40, 41, 52, 60, 61,63, 73, 75, 77, 79, 80, 81, and 88) is administered after treatment withan anti-CTLA4 antibody (e.g., ipilimumab) with or without a BRAFinhibitor (e.g., vemurafenib or dabrafenib).

In some embodiments, the immune checkpoint therapy is a costimulatoryligand. In some embodiments, the costimulatory ligand is OX40L, 41BBL,CD153, ICOSL, or CD40L.

In some embodiments, the immune checkpoint therapy is a MCSF/CSF-1Rinhibitor. An anti-CSF-1R can deplete TAMs, resulting in tumor growthinhibition. Cancer Cell 25, 1-14, Jun. 16, 2014. In some embodiments,the CSF-1R inhibitor is BLZ945, GW2850, R05509554, or PLX3397. In someembodiments, the CSF-1R inhibitor is BLZ945 or GW2850. In someembodiments, the CSF-1R inhibitor is PLX3397.

In some embodiments, the immune checkpoint therapy is animmunostimulant. In some embodiments, the immunostimulant is GMCSF, TLRligands, 41BBL, or ICOSL.

In some embodiments, the immune checkpoint therapy is a CXCR4/CXCL12inhibitor. In some embodiments, the CXCR4/CXCL12 inhibitor is AMD3100,AMD11070, AMD12118, AMD11814, or AMD13073. In some embodiments, theCXCR4/CXCL12 inhibitor is AMD3100.

In some embodiments, the immunotherapy is a CCL2 and/or CCR2 antagonist.In some embodiments, the antagonist of CCL2 and/or CCR2 is an anti-CCL2or CCR2 antibody. CCL2 is a chemokine and CCR2 is a chemokine receptor.CCL2 and CCR2, according to non-limiting theory, play a role in MDSCmigration.

In some embodiments, a PI3K-γ inhibitor disclosed herein, e.g., Compound4, is administered in combination with a BTK inhibitor. In oneembodiment, the BTK inhibitor is BTK inhibitors such as ibrutinib,AVL-292, Dasatinib, LFM-AI3, ONO-WG-307, and GDC-0834.

In some embodiments, a PI3K-γ inhibitor disclosed herein, e.g., Compound4, is administered in combination with an IDO (indoleamine2,3-dioxygenase) inhibitor or an TDO (tryptophan 2,3-dioxygenase)inhibitor. In one embodiment, the IDO inhibitor is indoximod, NLG919,INCB024360, F001287, norharmane, rosmarinic acid, oralpha-methyl-tryptophan. Although IDO inhibitors act within the TME,they do not specifically target MDSCs. The overexpression of IDO bydendritic cells creates an immunosuppressive tumor microenvironment.

In some embodiments, a PI3K-γ inhibitor disclosed herein, e.g., Compound4, is administered in combination with an inhibitor of one or moremembers of TAM family, a receptor tyrosine kinase (RTK) subfamilycomprising Tyro-3 (also called Sky), Axl and Mer. In one embodiment, theTAM inhibitor is BGB324 (R428), S49076, TP0903, CEP-40783, ONO-9330547,bosutinib (SKI606, PF5208763), cabozantinib (XL184), sunitinib(SU11248), foretinib (XL880, GSK1363089), MGCD265, BMS777607 (ASLAN002),LY2801653, SGI7079, amuvatinib (SGI-0470-02, MP470), SNS314,PF-02341066, diaminopyrimidine, spiroindoline, UNC569, UNC1062, UNC1666,UNC2025, or LDC1267. Additional TAM inhibitors include those describedin Mollard et al., Med. Chem. Lett. 2011, 2, 907-912 and Feneyrolles etal., Mol. Cancer Ther. 13(9), Published OnlineFirst Aug. 19, 2014, theentireties of which are incorporated by reference herein.

In some embodiment, a PI3K-γ inhibitor disclosed herein, e.g., Compound4, is administered to a subject concurrent or prior to theadministration of immune checkpoint therapy. In some embodiment, animmunostimulant is administered to a subject concurrent or prior to theadministration of immune checkpoint therapy. In some embodiment,chemotherapy (e.g., carboplatin, oxaliplatin, or radiation) isadministered to a subject concurrent or prior to the administration ofimmune checkpoint therapy.

In some embodiments, a PI3K-γ inhibitor disclosed herein, e.g., Compound4, is administered in combination with an ARG1 inhibitor. While notwishing to be bound by theory, it has been reported that tumorassociated myeloid cells establish an immunosuppressive microenvironmentin tumors through the expression of Arginase-1, which depletes the tumormicroenvironment of arginine, thereby the death or inhibition ofanti-tumor immune cells. Schmid et al., Proceedings: AACR 103rd AnnualMeeting 2012, Cancer Research: Apr. 15, 2012; Volume 72, Issue 8,Supplement 1. It has been reported that suppression of PI3Kgamma orArginase-1 expression blocked myeloid cell induced death of T cells invitro. Id. According to the non-limiting theory, PI3Kgamma inhibitionblocks Arginase-1 expression, thereby increasing the number of CD8+ Tcells in tumors, stimulating T cell-mediated cytotoxicity of tumorcells, and suppressing growth and metastasis of tumors. Combinationtherapies can be designed in accordance with this mechanism.

For instance, in some embodiments, a PI3K-γ inhibitor disclosed herein,e.g., Compound 4, is administered in combination with an ARG1 inhibitor.The ARG1 inhibitor may be, e.g., an inhibitory nucleic acid such as asiRNA, an inhibitory anti-ARG-1 antibody, or an analog of arginine.Other exemplary inhibitors of ARG1 include N-hydroxy-guanidinium orN-hydroxy-nor-1-arginine, and boronic acid derivatives, such as,2(S)-amino-6-boronohexanoic acid, and S-(2-boronoethyl)-1-cysteine,α-α-disubstituted amino acid based arginase inhibitors [such as(R)-2-amino-6-borono-2-(2-(piperidin-1-yl)ethyl)hexanoic acid], andpiceatannol-3′-O-β-d-glucopyranoside (PG). Steppan et al., “Developmentof novel arginase inhibitors for therapy of endothelial dysfunction.”,Front Immunol. 2013 Sep. 17; 4:278. doi: 10.3389/fimmu.2013.00278.

The PI3K γ inhibitors disclosed herein can have minimal effects onT-cell activation when compared to the suppressive effect of a PI3K δinhibitor on T-cell activation. Lewis lung carcinoma tumor growth can bereduced in PI3K γ knockout mice and can have decreased tumor associatedsuppressive myeloid cell infiltrates. Tumor associated suppressivemyeloid cells can include e.g., myeloid derived suppressor cells (MDSCs)and tumor associated macrophages (TAMs). PI3K γ knockout mice have TAMswhere the M2 phenotype is lost. M2 cells are immunosuppressive andsupport tumor growth. PI3K inhibitors provided herein can block M2phenotype (e.g., in an in vitro differentiation system), and thus canslow tumor growth.

For example, the effect of PI3K γ inhibitors and PI3K δ inhibitors on Tcell activation as measured by inhibition of IFN-γ in response to ConAhas shown that PI3K-δ is plays a role in mediating T cell activation,while PI3K-γ has minimal effects on T-cell activation. The IC₅₀ for aPI3K δ inhibitor in this assay is 3 nM, and the IC₅₀ for a PI3K γinhibitor is 2500 nM. Administration of PI3K-γ inhibitors can lead toimpaired T-cell migration but may have reduced effects on T-cellproliferation or activation.

In some embodiments, the PI3K γ inhibitors disclosed herein can havepotent effects on tumor associated suppressive myeloid cells withoutinhibiting the effector T-cell. The PI3K γ inhibitors disclosed hereincan have potent effects on tumor associated suppressive myeloid cellswithout blocking anti-tumor T-cell effects and thus can increase T cellactivity. In one embodiment, this effect can be enhanced byadministering CTLA4 antagonists and/or PD-1 and PDL1 antagonists. ThePI3K γ inhibitors disclosed herein can increase T cell activation andproliferation. In some embodiments, provided herein is a method ofblocking tumor associated suppressive myeloid cells without inhibitingthe effects on anti-tumor T-cells comprising administering an effectiveamount of a PI3K γ inhibitor disclosed herein or a pharmaceuticallyacceptable salt thereof to a subject. In some embodiments, providedherein is a method of blocking tumor associated suppressive myeloidcells without inhibiting the effects on anti-tumor T-cells comprisingadministering an effective amount of a compound disclosed herein or apharmaceutically acceptable salt thereof to a subject. In someembodiments, the subject has lung cancer, breast cancer, glioblastoma,or lymphoma (e.g., non-Hodgkin's lymphoma).

Further provided herein are methods of modulating kinase activity bycontacting a kinase with an amount of a compound provided hereinsufficient to modulate the activity of the kinase. Modulate can beinhibiting or activating kinase activity. In some embodiments, providedherein are methods of inhibiting kinase activity by contacting a kinasewith an amount of a compound provided herein sufficient to inhibit theactivity of the kinase. In some embodiments, provided herein are methodsof inhibiting kinase activity in a solution by contacting said solutionwith an amount of a compound provided herein sufficient to inhibit theactivity of the kinase in said solution. In some embodiments, providedherein are methods of inhibiting kinase activity in a cell by contactingsaid cell with an amount of a compound provided herein sufficient toinhibit the activity of the kinase in said cell. In some embodiments,provided herein are methods of inhibiting kinase activity in a tissue bycontacting said tissue with an amount of a compound provided hereinsufficient to inhibit the activity of the kinase in said tissue. In someembodiments, provided herein are methods of inhibiting kinase activityin an organism by contacting said organism with an amount of a compoundprovided herein sufficient to inhibit the activity of the kinase in saidorganism. In some embodiments, provided herein are methods of inhibitingkinase activity in an animal by contacting said animal with an amount ofa compound provided herein sufficient to inhibit the activity of thekinase in said animal. In some embodiments, provided herein are methodsof inhibiting kinase activity in a mammal by contacting said mammal withan amount of a compound provided herein sufficient to inhibit theactivity of the kinase in said mammal. In some embodiments, providedherein are methods of inhibiting kinase activity in a human bycontacting said human with an amount of a compound provided hereinsufficient to inhibit the activity of the kinase in said human. In someembodiments, the % of kinase activity after contacting a kinase with acompound provided herein is less than 1, 5, 10, 20, 30, 40, 50, 60, 70,80 90, 95, or 99% of the kinase activity in the absence of saidcontacting step.

In certain embodiments, provided herein are pharmaceutical compositioncomprising a therapeutically effective amount of a compound providedherein (e.g., a compound of any of Formulae (I″), (I′), (A′), (I), (A),(II), (III), (IV), (V), (VI), (VII), (IX), (X), (XI), (XII), (XIII),(XIV), (XV), (XVI), and (XVII), e.g., a selective PI3K-γ inhibitor,e.g., Compound 4), or a pharmaceutically acceptable form thereof, and animmunomodulator.

In one embodiment, the immunomodulator is an inhibitor of PD-1, PD-L1,LD-L2, CTLA-4, TIM3, LAG3, VISTA, BTLA, TIGIT, LAIR1, CD160, 2B4,TGFR-beta, or IDO/TDO, or a combination thereof. In one embodiment, theimmunomodulator is an inhibitor of PD-L1. In one embodiment, theimmunomodulator is an antibody or fragment thereof, an inhibitorynucleic acid, a soluble ligand, or a fusion of a PD-1 ligand with a Fcregion of an immunoglobulin. In one embodiment, the immunomodulator is acostimulatory ligand, a MCSF/CSF-1R inhibitor, an immunostimulant, aCXCR4/CXCL12 inhibitor, a CCL2 inhibitor, or a CCR2 inhibitor. In oneembodiment, the immunomodulator is cyclophosphamide, docetaxel,paclitaxel, 5-FU, or temozolomide.

In one embodiment, provided herein is a method of treating aPI3K-mediated disorder in a subject, comprising administering to thesubject a therapeutically effective amount of the composition.

Combination Therapy for Pulmonary and Respiratory Diseases

In some embodiments, the compound provided herein is administered incombination with one or more other therapies. Such therapies includetherapeutic agents as well as other medical interventions, behavioraltherapies (e.g., avoidance of sunlight), and the like.

By “in combination with,” it is not intended to imply that the othertherapy and the compound provided herein must be administered at thesame time and/or formulated for delivery together, although thesemethods of delivery are within the scope of this disclosure. Thecompound provided herein can be administered concurrently with, prior to(e.g., 5 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4hours, 6 hours, 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, 1week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, 12 weeks, or16 weeks before), or subsequent to (e.g., 5 minutes, 15 minutes, 30minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24hours, 48 hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks,5 weeks, 6 weeks, 8 weeks, 12 weeks, or 16 weeks after), one or moreother therapies (e.g., one or more other additional agents). In general,each therapeutic agent will be administered at a dose and/or on a timeschedule determined for that particular agent. The other therapeuticagent can be administered with the compound provided herein in a singlecomposition or separately in a different composition. Triple therapy isalso contemplated herein.

In general, it is expected that additional therapeutic agents employedin combination be utilized at levels that do not exceed the levels atwhich they are utilized individually. In some embodiments, the levelsutilized in combination will be lower than those utilized individually.

In some embodiments, the compound provided herein is a first linetreatment for a pulmonary or respiratory disease, i.e., it is used in asubject who has not been previously administered another drug intendedto treat a pulmonary or respiratory disease, or one or more symptoms ofthe disease.

In some embodiments, the compound provided herein is a second linetreatment for a pulmonary or respiratory disease, i.e., it is used in asubject who has been previously administered another drug intended totreat a pulmonary or respiratory disease, or one or more symptoms of thedisease.

In some embodiments, the compound provided herein is a third or fourthline treatment for a pulmonary or respiratory disease, i.e., it is usedin a subject who has been previously administered two or three otherdrugs intended to treat a pulmonary or respiratory disease, or one ormore symptoms of the disease.

In embodiments where two agents are administered, the agents can beadministered in any order. For example, the two agents can beadministered concurrently (i.e., essentially at the same time, or withinthe same treatment) or sequentially (i.e., one immediately following theother, or alternatively, with a gap in between administration of thetwo). In some embodiments, the compound provided herein is administeredsequentially (i.e., after the first therapeutic).

In some embodiments, the compound provided herein and the second agentare administered as separate compositions, e.g., pharmaceuticalcompositions. In some embodiments, the compound provided herein and theagent are administered separately, but via the same route (e.g., both byinhalation). In some embodiments, the compound provided herein and theagent are administered in the same composition, e.g., pharmaceuticalcomposition.

In some embodiments, the compound provided herein (e.g., PI3Kδ inhibitoror PI3Kγ inhibitor) is administered in combination with an agent thatinhibits IgE production or activity. In some embodiments, the compoundprovided herein (e.g., PI3Kδ inhibitor or PI3Kγ inhibitor) isadministered in combination with an inhibitor of mTOR. Agents thatinhibit IgE production are known in the art and they include but are notlimited to one or more of TEI-9874,2-(4-(6-cyclohexyloxy-2-naphtyloxy)phenylacetamide)benzoic acid,rapamycin, rapamycin analogs (i.e., rapalogs), TORC1 inhibitors, TORC2inhibitors, and any other compounds that inhibit mTORC1 and mTORC2.Agents that inhibit IgE activity include, for example, anti-IgEantibodies such as for example Omalizumab and TNX-901.

In certain embodiments, wherein inflammation (e.g., COPD, asthma) istreated, prevented and/or managed, a compound provided herein can becombined with, for example: PI3K inhibitors such as RP-6530, TG 100-115,RV1729, GS-1101, XL 499, GDC-0941, and AMG-319; BTK inhibitors such asibrutinib and AVL-292; JAK inhibitors such as tofacitinib and GLPG0636;SYK inhibitors such as fostamatinib.

In some embodiments, a compound provided herein can be combined withother agents that act to relieve the symptoms of inflammatoryconditions, such as COPD, asthma, and the other diseases describedherein. These agents include, but are not limited to, non-steroidalanti-inflammatory drugs (NSAIDs), e.g., acetylsalicylic acid; ibuprofen;naproxen; indomethacin; nabumetone; and tolmetin. In some embodiments,corticosteroids are used to reduce inflammation and suppress activity ofthe immune system.

In some embodiments, a compound provided herein (e.g., Compound 4), oran enantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof, isadministered in combination with an agent for pulmonary or respiratorydiseases. Examples of agents for pulmonary or respiratory diseasesinclude, but are not limited to, Abraxane (paclitaxel protein-boundparticles for injectable suspension), Adempas (riociguat), Anoro Ellipta(umeclidinium and vilanterol inhalation powder), Breo Ellipta(fluticasone furoate and vilanterol inhalation powder), Opsumit(macitentan), Qnasl (beclomethasone dipropionate) nasal aerosol, Sirturo(bedaquiline), Dymista (azelastine hydrochloride and fluticasonepropionate), Kalydeco (ivacaftor), Qnasl (beclomethasone dipropionate)nasal aerosol, Rayos (prednisone) delayed-release tablets, Surfaxin(lucinactant), Tudorza Pressair (aclidinium bromide inhalation powder),Arcapta (indacaterol maleate inhalation powder), Daliresp (roflumilast),Xalkori (crizotinib), Cayston (aztreonam for inhalation solution),Dulera (mometasone furoate+formoterol fumarate dihydrate), Teflaro(ceftaroline fosamil), Adcirca (tadalafil), Tyvaso (treprostinil),Alvesco (ciclesonide), Patanase (olopatadine hydrochloride), Letairis(ambrisentan), Xyzal (levocetirizine dihydrochloride), Brovana(arformoterol tartrate), Tygacil (tigecycline), Ketek (telithromycin),Spiriva HandiHaler (tiotropium bromide), Aldurazyme (laronidase), Iressa(gefitinib), Xolair (omalizumab), Zemaira (alpha1-proteinase inhibitor),Clarinex, Qvar (beclomethasone dipropionate), Remodulin (treprostinil),Xopenex (levalbuterol), Avelox I.V. (moxifloxacin hydrochloride), DuoNeb(albuterol sulfate and ipratropium bromide), Foradil Aerolizer(formoterol fumarate inhalation powder), Invanz, NasalCrom Nasal Spray,Tavist (clemastine fumarate), Tracleer (bosentan), Ventolin HFA(albuterol sulfate inhalation aerosol), Biaxin XL (clarithromycinextended-release tablets), Cefazolin and Dextrose USP, Tri-Nasal Spray(triamcinolone acetonide spray), Accolate (zafirlukast), CafcitInjection, Proventil HFA Inhalation Aerosol, Rhinocort Aqua Nasal Spray,Tequin, Tikosyn Capsules, Allegra-D, Clemastine fumarate syrup,Curosurf, Dynabac, Infasurf, Priftin, Pulmozyme (dornase alfa),Sclerosol Intrapleural Aerosol, Singulair (montelukast sodium), Synagis,Ceftin (cefuroxime axetil), Cipro (ciprofloxacin HCl), Claritin RediTabs(10 mg loratadine rapidly-disintegrating tablet), Flonase Nasal Spray,Flovent Rotadisk, Metaprotereol Sulfate Inhalation Solution (5%),Nasacort AQ (triamcinolone acetonide) Nasal Spray, Omnicef, Raxar(grepafloxacin), Serevent, Tilade (nedocromil sodium), Tobi, Vanceril 84mcg Double Strength (beclomethasone dipropionate, 84 mcg) InhalationAerosol, Zagam (sparfloxacin) tablets, Zyflo (Zileuton), Allegra(fexofenadine hydrochloride), Astelin nasal spray, Atrovent (ipratropiumbromide), Augmentin (amoxicillin/clavulanate), Azmacort (triamcinoloneacetonide) Inhalation Aerosol, Breathe Right, Claritin Syrup(loratadine), Claritin-D 24 Hour Extended Release Tablets (10 mgloratadine, 240 mg pseudoephedrine sulfate), Covera-HS (verapamil),OcuHist, RespiGam (Respiratory Syncitial Virus Immune GlobulinIntravenous), Tripedia (Diptheria and Tetanus Toxoids and AcellularPertussis Vaccine Absorbed), Vancenase AQ 84 mcg Double Strength,Visipaque (iodixanol), Zosyn (sterile piperacillin sodium/tazobactamsodium), Cedax (ceftibuten), and Zyrtec (cetirizine HCl). In oneembodiment, the agent for pulmonary or respiratory diseases is Arcapta(indacaterol maleate inhalation powder), Daliresp (roflumilast), Dulera(mometasone furoate+formoterol fumarate dihydrate), Alvesco(ciclesonide), Brovana (arformoterol tartrate), Spiriva HandiHaler(tiotropium bromide), Xolair (omalizumab), Qvar (beclomethasonedipropionate), Xopenex (levalbuterol), DuoNeb (albuterol sulfate andipratropium bromide), Foradil Aerolizer (formoterol fumarate inhalationpowder), Accolate (zafirlukast), Singulair (montelukast sodium), FloventRotadisk (fluticasone propionate inhalation powder), Tilade (nedocromilsodium), Vanceril (beclomethasone dipropionate, 84 mcg), Zyflo(Zileuton), and Azmacort (triamcinolone acetonide) Inhalation Aerosol.In one embodiment, the agent for pulmonary or respiratory diseases isSpiriva HandiHaler (tiotropium bromide).

Examples of agents for pulmonary or respiratory diseases include, butare not limited to, acetylcysteine (mucomyst) selected from TudorzaPressair (aclidinium bromide), Atrovent (ipratropium), andSpiriva_(tiotropium).

Examples of agents for pulmonary or respiratory diseases include, butare not limited to, beta2 agonists selected from short-acting beta2agonists and long acting beta2 agonists. Short acting beta2 agonistsinclude, but are not limited to, Proventil (albuterol), Tornalate(bitolterol), Xopenex (levalbuterol), Maxair (pirbuterol), and Alupent(metaproterenol). Long acting beta2 agonists include, but are notlimited to, Brovana (arformoterol tartrate), Foradil (formoterol),Arcapta Neohaler (indacaterol maleate), and Serevent (salmeterol).

Examples of agents for pulmonary or respiratory diseases include, butare not limited to, combination of two agents. In one embodiment, thecombination is administered through inhalation. The combination of twoagents includes, but is not limited to a beta2 agonist and ananticholinergi selected from Combivent (albuterol and ipratropium) andAnoro Ellipta (umeclidinium and vilanterol inhalation powder). Thecombination of two agents include, but are not limited to a beta2agonist and a corticosteroid selected from Advair (fluticasone andsalmeterol), Breo Ellipta (fluticasone furoate and vilanterol inhalationpowder), Dulera (mometasone furoate and formoterol fumarate), andSymbicort (budesonide and formoterol).

Examples of agents for pulmonary or respiratory diseases include, butare not limited to, corticosteroids selected from Vanceril Beclovent(beclomethasone), Pulmicort (budesonide), Alvesco (ciclesonide), Aerobid(flunisolide), Flovent (fluticasone), Asmanex (mometasone furoate), andAzmacort (triamcinolone).

Examples of agents for pulmonary or respiratory diseases include, butare not limited to, leukotriene inhibitors selected from Singulair(montelukast), Accolate (zafirlukast), and Zyflo (zileuton).

Examples of agents for pulmonary or respiratory diseases include, butare not limited to, mast cell stabilizers selected from Intal (cromolynsodium) and Tilade (nedocromil).

Examples of agents for pulmonary or respiratory diseases include, butare not limited to, phosphodiesterase 4 (PDE4) inhibitors selected fromDaliresp (roflumilast).

In some embodiments, a compound provided herein (e.g., Compound 4), oran enantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable form thereof, isadministered in combination with an agent for immunology or infectiousdiseases. Examples of agents for immunology or infectious diseasesinclude, but are not limited to, Kineret (anakinra), Lovenox (enoxaparinsodium) Injection, Makena (hydroxyprogesterone caproate injection),Myalept (metreleptin for injection), Qnasl (beclomethasone dipropionate)nasal aerosol, Simponi (golimumab), Sitavig (acyclovir) buccal tablets,Tecfidera (dimethyl fumarate), Tivicay (dolutegravir), VariZIG,Varicella Zoster Immune Globulin (Human), Flublok (seasonal influenzavaccine), Flucelvax (influenza virus vaccine), Fulyzaq (crofelemer),Horizant (gabapentin enacarbil), Qnasl (beclomethasone dipropionate)nasal aerosol, Rayos (prednisone) delayed-release tablets, Stribild(elvitegravir, cobicistat, emtricitabine, tenofovir disoproxilfumarate), Tudorza Pressair (aclidinium bromide inhalation powder),Arcapta (indacaterol maleate inhalation powder), Benlysta (belimumab),Complera (emtricitabine/rilpivirine/tenofovir disoproxil fumarate),Daliresp (roflumilast), Dificid (fidaxomicin), Edurant (rilpivirine),Firazyr (icatibant), Gralise (gabapentin), Incivek (telaprevir), Nulojix(belatacept), Victrelis (boceprevir), Cayston (aztreonam for inhalationsolution), Egrifta (tesamorelin for injection), Menveo (meningitisvaccine), Oravig (miconazole), Prevnar 13 (Pneumococcal 13-valentConjugate Vaccine), Teflaro (ceftaroline fosamil), Zortress(everolimus), Zymaxid (gatifloxacin ophthalmic solution), Bepreve(bepotastine besilate ophthalmic solution), Berinert (C1 EsteraseInhibitor (Human)), Besivance (besifloxacin ophthalmic suspension),Cervarix [Human Papillomavirus Bivalent (Types 16 and 18) Vaccine,Recombinant], Coartem (artemether/lumefantrine), Hiberix (Haemophilus bConjugate Vaccine; Tetanus Toxoid Conjugate), Ilaris (canakinumab),Ixiaro (Japanese Encephalitis Vaccine, Inactivated, Adsorbed), Kalbitor(ecallantide), Qutenza (capsaicin), Vibativ (telavancin), Zirgan(ganciclovir ophthalmic gel), Aptivus (tipranavir), Astepro (azelastinehydrochloride nasal spray), Cinryze (C1 Inhibitor (Human)), Intelence(etravirine), Moxatag (amoxicillin), Rotarix (Rotavirus Vaccine, Live,Oral), Tysabri (natalizumab), Viread (tenofovir disoproxil fumarate),Altabax (retapamulin), AzaSite (azithromycin), Doribax (doripenem),Extina (ketoconazole), Isentress (raltegravir), Selzentry (maraviroc),Veramyst (fluticasone furoate), Xyzal (levocetirizine dihydrochloride),Eraxis (anidulafungin), Gardasil (quadrivalent human papillomavirus(types 6, 11, 16, 18) recombinant vaccine), Noxafil (posaconazole),Prezista (darunavir), Rotateq (rotavirus vaccine, live oralpentavalent), Tyzeka (telbivudine), Veregen (kunecatechins), Baraclude(entecavir), Tygacil (tigecycline), Ketek (telithromycin), Tindamax,tinidazole, Xifaxan (rifaximin), Amevive (alefacept), FluMist (InfluenzaVirus Vaccine), Fuzeon (enfuvirtide), Lexiva (fosamprenavir calcium),Reyataz (atazanavir sulfate), Alinia (nitazoxanide), Clarinex, Daptacel,Fluzone Preservative-free, Hepsera (adefovir dipivoxil), PediarixVaccine, Pegasys (peginterferon alfa-2a), Restasis (cyclosporineophthalmic emulsion), Sustiva, Vfend (voriconazole), Avelox I.V.(moxifloxacin hydrochloride), Cancidas, Peg-Intron (peginterferonalfa-2b), Rebetol (ribavirin), Spectracef, Twinrix, Valcyte(valganciclovir HCl), Xigris (drotrecogin alfa [activated]), ABREVA(docosanol), Biaxin XL (clarithromycin extended-release tablets),Cefazolin and Dextrose USP, Children's Motrin Cold, Evoxac, KaletraCapsules and Oral Solution, Lamisil (terbinafine hydrochloride) Solution(1%), Lotrisone (clotrimazole/betamethasone diproprionate) lotion,Malarone (atovaquone; proguanil hydrochloride) Tablet, Rapamune(sirolimus) Tablets, Rid Mousse, Tri-Nasal Spray (triamcinoloneacetonide spray), Trivagizole 3 (clotrimazole) Vaginal Cream, Trizivir(abacavir sulfate; lamivudine; zidovudine AZT) Tablet, Agenerase(amprenavir), Cleocin (clindamycin phosphate), Famvir (famciclovir),Norvir (ritonavir), Panretin Gel, Rapamune (sirolimus) oral solution,Relenza, Synercid I.V., Tamiflu capsule, Vistide (cidofovir), Allegra-D,CellCept, Clemastine fumarate syrup, Dynabac, REBETRON™ CombinationTherapy, Simulect, Timentin, Viroptic, INFANRIX (Diphtheria and TetanusToxoids and Acellular Pertussis Vaccine Adsorbed), Acyclovir Capsules,Aldara (imiquimod), Aphthasol, Combivir, Condylox Gel 0.5% (pokofilox),Flagyl ER, Flonase Nasal Spray, Fortovase, INFERGEN (interferonalfacon-1), Intron A (interferon alfa-2b, recombinant), RescriptorTablets (delavirdine mesylate tablets), SPORANOX (itraconazole),Stromectol (ivermectin), Taxol, Trovan, VIRACEPT (nelfinavir mesylate),Zerit (stavudine), Albenza (albendazole), Apthasol (Amlexanox),Carrington patch, Confide, Crixivan (Indinavir sulfate), Gastrocrom OralConcentrate (cromolyn sodium), Havrix, Lamisil (terbinafinehydrochloride) Tablets, Leukine (sargramostim), Oral Cytovene, RespiGam(Respiratory Syncitial Virus Immune Globulin Intravenous), Videx(didanosine), Viramune (nevirapine), Vitrasert Implant, Zithromax(azithromycin), Cedax (ceftibuten), Clarithromycin (Biaxin), Epivir(lamivudine), Invirase (saquinavir), Valtrex (valacyclovir HCl), andZyrtec (cetirizine HCl).

The examples and preparations provided below further illustrate andexemplify the compounds as provided herein and methods of preparing suchcompounds. It is to be understood that the scope of the presentdisclosure is not limited in any way by the scope of the followingexamples and preparations. In the following examples molecules with asingle chiral center, unless otherwise noted, exist as a racemicmixture. Those molecules with two or more chiral centers, unlessotherwise noted, exist as a racemic mixture of diastereomers. Singleenantiomers/diastereomers can be obtained by methods known to thoseskilled in the art.

Synthesis of Compounds

In some embodiments, compounds of provided herein may be preparedaccording to methods known in the art. For example, the compoundsprovided herein can be synthesized according to the schemes below.Scheme 1, shows the synthesis of amine A-30, F-50, X-40, and H50. Scheme2 shows the synthesis of amide D-20 and formula I.

Specifically, in Scheme 1 in method A, isoquinolinone amine compoundA-30 is generated in two steps. For example, in the first step, compoundA-10 is converted to compound A-20. Compound A-20 is coupled withtert-butyl (1-(methoxy(methyl)amino)-1-oxopropan-2-yl)carbamate toafford compound A-30. In some embodiments, isoquinolinone compounds canbe prepared according to method H. For example, compound H-10 is coupledwith tert-butyl (1-(methoxy(methyl)amino)-1-oxopropan-2-yl)carbamate togenerate compound H-20, which is then converted to H-30. Compound H-30is reacted with B—NH₂ to form compound H-40, which is then treated withe.g., an acid to afford H-50.

In method F, quinazolinone F-50 is generated. For example, compound F-10is converted to compound F-20, which couples with2-((tert-butoxycarbonyl)amino)propanoic acid to form F-30. Compound F-30is then converted to F-40. Compound F-40 is deprotected to affordcompound F-50. Alternatively, quinazolinone X-40 can be preparedstarting with 2-amino-6-chlorobenzoic acid to generate compound X-10,which may be converted to compound X-20. Compound X-20 may be coupledwith 2-((tert-Butoxycarbonyl)amino)propanoic acid to generate compoundX-30, which may be converted to the desired compound X-40.

In Scheme 2, amine compound A30, F50, X-40, or H50 is treated withWd-C(O)OH to afford amide D20, which is treated with an alkyne togenerate a compound of Formula (I).

EXAMPLES Chemical Examples

The chemical entities described herein can be synthesized according toone or more illustrative schemes herein and/or techniques well known inthe art.

Unless specified to the contrary, the reactions described herein takeplace at atmospheric pressure, generally within a temperature range from−10° C. to 200° C. Further, except as otherwise specified, reactiontimes and conditions are intended to be approximate, e.g., taking placeat about atmospheric pressure within a temperature range of about −10°C. to about 110° C. over a period that is, for example, about 1 to about24 hours; reactions left to run overnight in some embodiments canaverage a period of about 16 hours.

The terms “solvent,” “organic solvent,” and “inert solvent” each mean asolvent inert under the conditions of the reaction being described inconjunction therewith including, for example, benzene, toluene,acetonitrile, tetrahydrofuran (“THF”), dimethylformamide (“DMF”),chloroform, methylene chloride (or dichloromethane), diethyl ether,methanol, N-methylpyrrolidone (“NMP”), pyridine, and the like. Unlessspecified to the contrary, the solvents used in the reactions describedherein are inert organic solvents. Unless specified to the contrary, foreach gram of the limiting reagent, one cc (or mL) of solvent constitutesa volume equivalent.

Isolation and purification of the chemical entities and intermediatesdescribed herein can be effected, if desired, by any suitable separationor purification procedure, such as, for example, filtration, extraction,crystallization, column chromatography, thin-layer chromatography, orthick-layer chromatography, or a combination of these procedures.Specific illustrations of suitable separation and isolation proceduresare given by reference to the examples herein below. However, otherequivalent separation or isolation procedures can also be used.

When desired, the (R)- and (S)-isomers of the non-limiting exemplarycompounds, if present, can be resolved by methods known to those skilledin the art, for example by formation of diastereoisomeric salts orcomplexes which can be separated, for example, by crystallization; viaformation of diastereoisomeric derivatives which can be separated, forexample, by crystallization, gas-liquid or liquid chromatography;selective reaction of one enantiomer with an enantiomer-specificreagent, for example enzymatic oxidation or reduction, followed byseparation of the modified and unmodified enantiomers; or gas-liquid orliquid chromatography in a chiral environment, for example on a chiralsupport, such as silica with a bound chiral ligand or in the presence ofa chiral solvent. Alternatively, a specific enantiomer can besynthesized by asymmetric synthesis using optically active reagents,substrates, catalysts or solvents, or by converting one enantiomer tothe other by asymmetric transformation. Further, atropisomers (i.e.,stereoisomers from hindered rotation about single bonds) of compoundsprovided herein can be resolved or isolated by methods known to thoseskilled in the art. For example, certain B substituents with ortho ormeta substituted phenyl may form atropisomers, where they may beseparated and isolated.

The compounds described herein can be optionally contacted with apharmaceutically acceptable acid to form the corresponding acid additionsalts. Also, the compounds described herein can be optionally contactedwith a pharmaceutically acceptable base to form the corresponding basicaddition salts.

In some embodiments, compounds provided herein can generally besynthesized by an appropriate combination of generally well knownsynthetic methods. Techniques useful in synthesizing these chemicalentities are both readily apparent and accessible to those of skill inthe relevant art, based on the instant disclosure. Many of theoptionally substituted starting compounds and other reactants arecommercially available, e.g., from Aldrich Chemical Company (Milwaukee,Wis.) or can be readily prepared by those skilled in the art usingcommonly employed synthetic methodology.

The discussion below is offered to illustrate certain of the diversemethods available for use in making the compounds and is not intended tolimit the scope of reactions or reaction sequences that can be used inpreparing the compounds provided herein.

General Synthetic Methods

The compounds herein being generally described, it will be more readilyunderstood by reference to the following examples, which are includedmerely for purposes of illustration of certain aspects and embodiments,and are not intended to limit these aspects and embodiments.

(i) General Method for the Synthesis of Amine Cores:

Method A General Conditions for the Preparation of(S)-3-(1-aminoethyl)-isoquinolin-1(2H)-ones

To a stirred mixture of a given o-methylbenzoic acid (A-1) (1 eq, e.g.,1.5 mol) and DMF (catalytic, e.g., 2 mL) in DCM (1.2 M, e.g., 1275 mL)at RT, oxalyl chloride (1.1 eq, e.g., 1.65 mol) is added over 5 min andthe resulting mixture is stirred at RT for 2 h. The mixture is thenconcentrated in vacuo. The residue is dissolved in DCM (150 mL) and theresulting solution (solution A) is used directly in the next step.

To a stirred mixture of aniline (1.05 eq, e.g., 1.58 mol) andtriethylamine (2.1 eq, e.g., 3.15 mol) in DCM (1.2 M, e.g., 1350 mL),the above solution A (e.g., 150 mL) is added dropwise while the reactiontemperature is maintained between 25° C. to 40° C. by an ice-water bath.The resulting mixture is stirred at RT for 2 h and then water (e.g.,1000 mL) is added. The organic layers are separated and washed withwater (2× e.g., 1000 mL), dried over Na₂SO₄ and filtered. The filtrateis concentrated in vacuo. The product is suspended in n-heptanes (e.g.,1000 mL) and stirred at RT for 30 min. The precipitate is collected byfiltration, rinsed with heptanes (e.g., 500 mL) and further dried invacuo to afford the amide (A-2).

To a stirred mixture of amide (A-2) (1 eq, e.g., 173 mmol) in anhydrousTHF (e.g., 250 mL) at −30° C. under an argon atmosphere, a solution ofn-butyllithium in hexanes (2.5 eq, 2.5 M, e.g., 432 mol) is addeddropwise over 30 min while keeping the inner temperature between −30° C.and −10° C. The resulting mixture is then stirred at −30° C. for 30 min.

To a stirred mixture of (S)-tert-butyl1-(methoxy(methyl)amino)-1-oxopropan-2-ylcarbamate (1.5 eq, e.g., 260mmol) in anhydrous THF (e.g., 250 mL) at −30° C. under an argonatmosphere, a solution of isopropylmagnesium chloride in THF (1.65 eq, 1M, e.g., 286 mmol) is added dropwise over 30 min while keeping innertemperature between −30° C. and −10° C. The resulting mixture is stirredat −30° C. for 30 min. This solution is then slowly added to abovereaction mixture while keeping inner temperature between −30° C. and−10° C. The resulting mixture is stirred at −15° C. for 1 h. Thereaction mixture is quenched with water (e.g., 50 mL) and then acidifiedwith conc. HCl at −10° C. to 0° C. to adjust the pH to 1-3. The mixtureis allowed to warm to RT and concentrated in vacuo. The residue isdissolved in MeOH (e.g., 480 mL), and then conc. HCl (e.g., 240 mL) isadded quickly at RT. The resulting mixture is stirred at reflux for 1 h.The reaction mixture is concentrated in vacuo to reduce the volume toabout 450 mL. The residue is extracted with a 2:1 mixture of heptane andethyl acetate (e.g., 2×500 mL). The aqueous layer is basified withconcentrated ammonium hydroxide to adjust the pH value to 9-10 whilekeeping the inner temperature between −10° C. and 0° C. The mixture isthen extracted with DCM (e.g., 3×300 mL), washed with brine, dried overMgSO₄ and filtered. The filtrate is concentrated in vacuo and theresidue is dissolved in MeOH (e.g., 1200 mL) at RT. To this solution,D-(−)-tartaric acid (0.8 eq, e.g., 21 g, 140 mmol) is added in oneportion at RT. After stirring at RT for 30 min, a white solidprecipitates and the mixture is slurried at RT for 10 h. The solid iscollected by filtration and rinsed with MeOH (e.g., 3×50 mL). Thecollected solid is suspended in water (e.g., 500 mL) and thenneutralized with concentrated ammonium hydroxide solution at RT toadjust the pH to 9-10. The mixture is extracted with DCM (e.g., 3×200mL). The combined organic layers are washed with brine, dried over MgSO₄and filtered. The filtrate is concentrated in vacuo to afford the(S)-3-(1-aminoethyl)-isoquinolin-1(2H)-ones (A-3).

Method B General Conditions for the Preparation of3-(aminomethyl)-isoquinolin-1(2H)-ones

A mixture of benzoic acid (B-1) (1 eq, e.g., 400 mmol), oxalyl chloride(2 eq, e.g., 101 g, 800 mmol) and DMF (catalytic, e.g., 0.2 ml) in DCM(1M, e.g., 400 mL) is stirred at RT for 2 h. The mixture is concentratedin vacuo to afford the acid chloride (B-2). The product obtained is useddirectly in the next step without further purification.

A mixture of R₂NH₂ amine (1.05 eq, e.g., 420 mmol) and triethylamine(1.7, e.g., 700 mmol) in DCM (1.4 M, e.g., 300 mL) is stirred at RT for10 min. To this mixture, acid chloride (B-2) (1 eq, e.g., 400 mmol) isadded dropwise, and the resulting mixture is stirred at RT for 30 min.The reaction mixture is poured into water (e.g., 300 mL) and extractedwith DCM (e.g., 3×200 mL), dried over anhydrous Na₂SO₄ and filtered. Thefiltrate is concentrated in vacuo to afford the product. The product issuspended in isopropyl ether (e.g., 300 mL), stirred at reflux for 30min, and then cooled to 0-5° C. The precipitate is collected byfiltration and further dried in vacuo to afford the product amide (B-3).

To a stirred solution of amide (B-3) (1.0 eq, e.g., 0.1 mol) inanhydrous THF (0.4 M, e.g., 225 mL) at −78° C. under an argonatmosphere, a solution of n-butyllithium in hexanes (2.5 M, 3 eq, e.g.,120 mL, 0.3 mol) is added dropwise over 1 h period of time while keepinginner temperature between −78° C. to −50° C. The resulting mixture isstirred at −70° C. for 1 h, and then diethyl oxalate (1.2 eq, e.g., 17.5g, 0.12 mol) is quickly added (with an increase in temperature to −20°C. upon addition). The mixture is stirred at −50° C. for 10 min, andthen quenched with water (e.g., 100 mL). The inorganic salt is removedby filtration, and the filtrate is washed with ethyl acetate (e.g.,2×100 mL). The combined organic layers are washed with brine (e.g., 100mL), dried over MgSO₄ and filtered. The filtrate is concentrated invacuo to afford the product as a semi-solid. The product is slurried inisopropyl ether (e.g., 100 mL) at RT for 10 min. The solid is collectedby filtration and further dried in vacuo to afford the product (B-4).The product obtained is used directly in the next step.

Compound (B-4) (1 eq, e.g., 88 mmol) is dissolved at 0.9 M with HCl/MeOH(100 mL, e.g., 10 M), and the resulting mixture is stirred at reflux for1 h. The reaction mixture is concentrated in vacuo, and the residue isslurried in ethyl acetate (100 mL) at RT for 30 min. The solid iscollected by filtration, rinsed with ethyl acetate (3×50 mL), andfurther dried in vacuo to afford the product (B-5).

To a stirred suspension of lithium aluminum hydride (3 eq., e.g., 15.6g, 410 mmol) in anhydrous THF (0.3 M, e.g., 500 mL) at −78° C. under anitrogen atmosphere, (B-5) (1 eq, e.g., 137 mmol) is slowly added over a10 min period of time. The resulting mixture is allowed to warm to −30°C. and stirred for 30 min. The mixture is then cooled to −78° C., andquenched carefully with water (e.g., 100 mL). The mixture is allowed towarm to RT, filtered through silica gel (e.g., 20 g), and the filtrateis concentrated in vacuo. The product mixture is poured into H₂O (e.g.,200 mL) and extracted with ethyl acetate (e.g., 3×200 mL). The combinedorganic layers are washed with brine (e.g., 100 mL), dried over Na₂SO₄and filtered. The filtrate is concentrated in vacuo. The product issuspended in ethyl acetate (e.g., 30 mL) and stirred for 10 min. Thesolid is collected by filtration and further dried in vacuo to affordthe product (B-6).

Phosphorus tribromide (1.2 eq, e.g., 3.42 g, 12.6 mmol) and DMF (2.0 eq,e.g., 1.6 g, 21.0 mmol) is dissolved in CH₃CN (0.13 M, e.g., 100 mL) andthe resulting mixture is stirred at −10° C. for 10 min. To this mixture,alcohol (B-6) (1.0 eq, 10.5 mmol) is added in portions. The resultingmixture is allowed to warm to RT and stirred for an additional 30 min.The reaction mixture is neutralized with saturated aqueous NaHCO₃solution at 0-5° C. and then filtered. The filtrate is extracted withethyl acetate (e.g., 3×100 mL). The combined organic layers are washedwith brine, dried over Na₂SO₄ and filtered. The filtrate is concentratedin vacuo and the residue is purified by flash column chromatography onsilica gel (20% ethyl acetate-petroleum ether) to afford the productbromide (B-7).

To a stirred mixture of phthalimide (1.1 eq, e.g., 6.93 mmol) in DMF(e.g., 20 mL) at RT, potassium-tert-butoxide (1.5 eq, e.g., 1.1 g, 9.45mmol) is added in portions over 10 min and then bromide (B-7) (1.0 eq,e.g., 6.3 mmol) is added. The resulting mixture is stirred at 100° C.for 2 h. The reaction mixture is allowed to cool to RT and then pouredinto ice-water (e.g., 30 mL). The mixture is extracted with ethylacetate (e.g., 3×20 mL). The combined organic layers are washed withbrine, dried over Na₂SO₄ and filtered. The filtrate is concentrated invacuo and the residue is purified by flash column chromatography onsilica gel (e.g., 16% ethyl acetate-petroleum ether) to afford theproduct dione (B-8).

Dione (B-8) (1.0 eq, e.g., 1.5 mmol) and hydrazine hydrate (e.g., 8.0eq, 600 mg, 12 mmol) are dissolved in EtOH (e.g., 20 mL) and theresulting mixture is stirred at reflux for 1 h. The mixture is allowedto cool to RT and then filtered. The filter cake is washed with EtOH(e.g., 10 mL). The combined filtrate is concentrated in vacuo and theresidue is purified by flash column chromatography on silica gel (e.g.,2.5% MeOH-DCM) to afford the amine (B-9).

(ii) General Methods for Amide Synthesis:

Method D

To a mixture of amine (D-1) (1.0 eq, e.g., 0.5 mmol), W_(d)—COOHcarboxylic acid (1.1 eq, e.g., 0.55 mmol), and N,N-diisopropylethylamine(2.0 eq, e.g., 0.17 mL, 1.0 mmol) in anhydrous DMF (e.g., 5 mL),1-hydroxybenzotriazole hydrate (1.3 eq, e.g., 0.65 mmol) and EDChydrochloride (1.3 eq, e.g., 0.65 mmol) are added sequentially and theresulting mixture is stirred at RT for 2-16 h. Ice-water or saturatedsodium carbonate solution is added to the reaction mixture and thenstirred for 10 min. The precipitate is collected by filtration, rinsedwith water and dried in vacuo. The solid collected is further purifiedby flash column chromatography on silica gel (e.g., 0-10% MeOH-DCM) toafford the product amide (D-2).

Method E

A solution of amine (D-1) (1 eq, e.g., 0.25 mmol), W_(d)—COOH carboxylicacid (1.1 eq), and 1-hydroxybenzotriazole hydrate (1.3 eq) indimethylformamide (0.1 M) is treated with diisopropylethylamine (2 eq)and then EDC hydrochloride (1.3 eq, e.g., 63 mg). The reaction mixtureis stirred at ambient temperature overnight. The reaction mixture isdiluted with water (5× solvent) and acetic acid (1.5 eq) is added, thenthe mixture is stirred in an ice bath for 40 min. The resultingprecipitate is collected by filtration, and washed with water (e.g., 3×3mL). The collected solid is dried in vacuo to afford amide (D-2).

Method F

To a stirred mixture of nitrobenzoic acid (F-1) (1.0 eq, 1.0 mol) andDMF (e.g., 2.0 mL) in toluene (e.g., 800 mL), thionyl chloride (4.0 eq,e.g., 292 mL, 1.0 mol) is added dropwise (over 15 min) and the resultingmixture is stirred at reflux for 1.5 h. The mixture is allowed to coolto RT and then concentrated in vacuo. The residue is dissolved in DCM(e.g., 100 mL) to form solution A, which is used directly in the nextstep.

To a stirred mixture of a given amine R₂—NH₂ (1.1 eq, e.g., 102.4 g, 1.1mol) and triethylamine (2.0 eq, e.g., 280 mL, 2.0 mol) in DCM (1.6 M,e.g., 700 mL), solution A is added dropwise while keeping the reactiontemperature below 10° C. The resulting mixture is allowed to warm to RTand then stirred at RT overnight. The reaction mixture is diluted withice-water (e.g., 1.0 L) and stirred for 15 min. The precipitate iscollected by filtration, rinsed with isopropyl ether (e.g., 3×100 mL)and petroleum ether (e.g., 3×100 mL), and then dried in vacuo to affordproduct amide (F-2).

A mixture of nitro-benzamide (F-2) (1.0 eq, e.g., 20.0 mmol) and DMF(cat.) in toluene (0.3 M, e.g., 60 mL) at RT, thionyl chloride (8.2 eq,e.g., 12 mL, 164 mmol) is added dropwise (over 5 min) and the resultingmixture is stirred at reflux for 2 h. The mixture is allowed to cool toRT and then concentrated in vacuo. The residue is dissolved in DCM(e.g., 10 mL) to form solution B, which is used directly in the nextstep.

To a stirred mixture of N-(tert-butoxycarbonyl)-L-alanine (0.8 eq, e.g.,16.0 mmol) and N,N-diisopropylethylamine (1.5 eq, e.g., 4.0 g, 31.0 mol)in DCM (0.8 M, e.g., 20 mL), solution B is added dropwise while keepingthe reaction temperature between 0-10° C. The resulting mixture isstirred at this temperature for 1 h and then stirred at RT overnight.The reaction mixture is quenched with ice-water (e.g., 100 mL). Theorganic layer is separated and the aqueous layer is extracted with DCM(e.g., 2×80 mL). The combined organic layers are washed with brine,dried over Na₂SO₄ and filtered. The filtrate is concentrated in vacuoand the residue is slurried in isopropyl ether (e.g., 100 mL) for 15min. The solid is collected by filtration and dried in vacuo to affordproduct (F-3).

To a suspension of zinc dust (10.0 eq, e.g., 7.2 g, 110 mmol) in glacialacetic acid (2.8 M, e.g., 40 mL) at 15° C., a solution of (F-3) (1.0 eq,e.g., 11.0 mmol) in glacial acetic acid (0.3 M, e.g., 40 mL) is addedand the resulting mixture is stirred at RT for 4 h. The mixture ispoured into ice-water (e.g., 200 mL) and neutralized with saturatedaqueous NaHCO₃ solution to adjust the pH to 8. The resulting mixture isextracted with DCM (e.g., 3×150 mL). The combined organic layers arewashed with brine, dried over Na₂SO₄ and filtered. The filtrate isconcentrated in vacuo and the residue is purified by flashchromatography on silica gel (7% ethyl acetate-petroleum ether) toafford product (F-4).

Compound (F-4) (1.0 eq, e.g., 0.5 mmol) is dissolved in hydrochloricmethanol solution (8 eq, e.g., 2N, 20 mL) and the resulting mixture isstirred at RT for 2 h. The mixture is concentrated in vacuo. The residueis diluted with water (30 mL) and then neutralized with saturatedaqueous NaHCO₃ to adjust the pH to 8 while keeping the temperature below5° C. The resulting mixture is extracted with DCM (e.g., 3×30 mL). Thecombined organic layers are washed with brine, dried over Na₂SO₄ andfiltered. The filtrate is concentrated in vacuo and the residue isslurried in petroleum ether (e.g., 10 mL). The solid is collected byfiltration and dried in vacuo to afford product (F-5).

The quinazolinone (F-5) can be used to synthesize compounds describedherein using, for example, Method D to couple the amine to W_(d) groups.

Method FF

Alternatively, compounds with a quinazolinone core can be preparedaccording to the procedures in PCT publication no. WO2013082540.

In Method FF, 2-Amino-6-chlorobenzoic acid (63 mmol, 1.0 equiv) isdissolved in acetonitrile (60 mL) in a 250 mL round bottomed-flask,placed under an atmosphere of Ar and heated to 50° C. Pyridine (2.0equiv) is added followed by dropwise the addition of a solution oftriphosgene (0.34 equiv in 30 mL acetonitrile) while maintaining theinternal temperature below 60° C. The mixture is then stirred at 50° C.for 2 h after which the solvent is removed under vacuum. The remainingresidue is dispersed in 50 mL of water and filtered. The resulting solidis washed with a minimal amount of acetonitrile to remove discolorationand then dried to provide desired anhydride X-1.

Anhydride X-1 (25.5 mmol, 1.0 equiv) is suspended in dioxane (40 mL)under an atmosphere of Ar in a 200 mL round bottomed-flask. Aniline (1.0equiv) is added dropwise. Heating is started at 40° C. and graduallyincreased to 100° C. After 4 h, the majority of starting material isconsumed after which the reaction is allowed to cool. The solvent isthen removed under vacuum to provide an oil which is redissolved intoluene followed by the addition of hexanes until the solvent appearsclose to partitioning. The mixture is stirred for 14 h after which asolid appeared in the flask. This solid is isolated via vacuumfiltration and washed with hexanes to provide the desired amide X-2 inhigh yield.

(S)-2-((tert-Butoxycarbonyl)amino)propanoic acid (33.0 mmol, 2.0 equiv)is dissolved in dry tetrahydrofuran (70 mL) under an atmosphere of Arafter which N-methylmorpholine (2.2 equiv) is added dropwise. Themixture is then cooled to −17° C. in an acetone/dry ice bath after whicha solution of isobutyl chloroformate (2.0 equiv in 10 mL of drytetrahydrofuran) is added dropwise to the mixture followed by stirringfor 30 min. A solution of amine X-2 (10 equiv in 10 mL of drytetrahydrofuran) is then added. The dry ice bath is then removed and themixture is stirred at RT for 90 min. It is then heated to 60° C. foranother 2 h after which it is allowed to cool. MTBE (150 mL) and water(150 mL) are then successively added with strong stirring. The phasesare separated and the organic phase is washed with water (2×50 mL) andbrine (50 mL) and dried over sodium sulfate. The solution is thenconcentrated under reduced pressure and the crude reside is purifiedusing flash silica gel chromatography (gradient 5-30 ethylacetate/hexanes) X-3 as the coupled product.

Compound X-3 (4.9 mmol, 1.0 equiv) is then suspended in acetonitrile(100 mL). Triethylamine (48 equiv) is then added with stirring followedby the dropwise addition of chlorotrimethylsilane (15 equiv). The flaskis then sealed and heated to 90° C. for 3d. The reaction is allowed tocool after which the solvent is removed under vacuum. The residue isthen dissolved in ethyl acetate (120 mL) and successively washed withsaturated sodium carbonate (1×100 mL), water (1×100 mL) and brine (1×100mL). The organic layer is then dried over anhydrous sodium sulfate andconcentrated under reduced pressure to provide cyclized product X-4. Theproduct can either be used directly in subsequent reactions or purifiedusing flash silica gel chromatography.

Method G General Conditions for the Preparation of(S)-3-(1-aminoethyl)1-8-(trifluoromethyl)isoquinolin-1(2H)-ones

To a suspension of 2-amino-6-methylbenzoic acid (G-1) (20.0 g, 132.0mmol, 1.0 eq) in H₂O (55 mL) at 0-5° C., conc. HCl (36.5%, 64 mL, 749mmol, 5.7 eq) is added slowly. After stirring for 15 min, the mixture isadded dropwise to a solution of sodium nitrite (12.02 g, 174.0 mmol,1.32 eq) in H₂O (36 mL) at 0-5° C., and the resulting mixture is stirredfor 1 h. The resulting solution is then added to a solution of KI (60.5g, 364.5 mmol, 2.76 eq) in H₂O (150 mL) at 0-5° C. The reaction mixtureis allowed to warm to RT and stirred at RT overnight. The mixture isextracted with ethyl acetate (3×100 mL). The combined organic layers arewashed with water (2×100 mL), dried over anhydrous Na₂SO₄ and filtered.The filtrate is concentrated in vacuo and the residue is purified byflash column chromatography on silica gel (0-20% ethyl acetate-petroether) to afford the product, 2-iodo-6-methylbenzoic acid (G-2).

To a stirred mixture of 2-iodo-6-methylbenzoic acid (G-2) (305.3 mmol,1.0 eq) and DMF (0.3 mL) in DCM (350 mL) at RT, oxalyl chloride (466.4mmol, 1.5 eq) is added dropwise. The resulting mixture is stirred at RTfor 3 h and then concentrated in vacuo. The residue is dissolved in DCM(50 mL) and the resulting solution (solution A) is used directly in thenext step.

To a stirred mixture of R3-substituted aniline (335.7 mmol, 1.1 eq) andtriethylamine (915.0 mmol, 3.0 eq) in DCM (350 mL), solution A (150 mL)is added dropwise while the reaction temperature is controlled below 30°C. by an ice-water bath. The reaction mixture is stirred at RT for 1 hand then quenched with water (200 mL). The organic layer is separated,washed with water (2×200 mL), dried over anhydrous Na₂SO₄ and filtered.The filtrate is concentrated in vacuo. The product is rinsed withisopropyl ether and dried in vacuo to afford the product amide (G-3).

A mixture of amide (G-3) (18.0 mmol, 1.0 eq), methyl2,2-difluoro-2-(fluorosulfonyl)acetate (72.9 mmol, 4.0 eq) and CuI (3.63mmol, 0.2 eq) in DMF (130 mL) is stirred at 70° C. under an argonatmosphere overnight. The mixture is allowed to cool to RT and thenconcentrated in vacuo to remove the solvent. The resulting residue ispartitioned between ethyl acetate (60 mL) and water (60 mL), and theaqueous layer is extracted with ethyl acetate (2×60 mL). The combinedorganic layers are washed with water (2×60 mL), dried over anhydrousNa₂SO₄ and filtered. The filtrate is concentrated in vacuo and theresidue is purified by flash column chromatography on silica gel toafford the product, trifluoromethyl amide (G-4).

To a stirred mixture of amide (G-4) (10.1 mmol, 1.0 eq) in anhydrous THF(25 mL) at −40° C. under an argon atmosphere, a solution ofn-butyllithium in THF (2.5 M, 25.3 mmol, 2.5 eq) is added dropwise (over15 min) and the inner temperature is controlled between −30° C. and −20°C. during the addition. The resulting mixture is stirred at −30° C. foran additional 1 h. To a stirred mixture of (S)-tert-butyl1-(methoxy(methyl)amino)-1-oxopropan-2-ylcarbamate (11.1 mmol, 1.1 eq)in anhydrous THF (20 mL) at −30° C. under an argon atmosphere, asolution of isopropylmagnesium chloride in THF (12.6 mmol, 1.25 eq) isadded dropwise (over 15 min) and the inner temperature is controlledbelow −20° C. during the addition. The resulting mixture is stirred at−15° C. for 1 h. This solution is then slowly added to above reactionmixture at −30° C. (over 10 min), and the resulting mixture is stirredat −30° C. for an additional 30 min. The reaction mixture is quenchedwith water (50 mL) and then acidified with conc. HCl at −5° C. to adjustthe pH to 5. The mixture is allowed to warm to RT and concentrated invacuo. The residue is dissolved in MeOH (10 mL), and then conc. HCl (10mL) is added quickly at RT. The resulting mixture is stirred at refluxfor 2 h, cooled to RT and then concentrated in vacuo. The residue issuspended in water (15 mL), basified with concentrated ammoniumhydroxide to adjust the pH to 9-10 while keeping the inner temperaturebelow 5° C. and then extracted with DCM (3×15 mL). The combined organiclayers are washed with brine, dried over MgSO₄ and filtered. Thefiltrate is concentrated in vacuo and the residue is dissolved in MeOH(70 mL). To this solution, D-(−)-tartaric acid (8.1 mmol, 0.8 eq) isadded in one portion at RT. After stirring at RT for 30 min, a solidprecipitates and the mixture is slurried at RT for 10 h. The precipitateis collected by filtration and rinsed with MeOH (3×4.0 mL). Thecollected solid is suspended in water (30 mL) and then neutralized withconcentrated ammonium hydroxide solution at RT to adjust the pH to 9-10.The mixture is extracted with DCM (3×15 mL). The combined organic layersare washed with brine, dried over anhydrous MgSO₄ and filtered. Thefiltrate is concentrated in vacuo to afford the product,(S)-3-(1-aminoethyl)-8-(trifluoromethyl)isoquinolin-1(2H)-one (G-5).

Method H General Conditions for the Preparation of(S)-3-(1-aminoethyl)-isoquinolin-1(2H)-ones

An o-methylbenzoic acid (H-1) (1 eq, e.g., 46.9 mmol) in a flame-driedround bottom flask under nitrogen is dissolved in THF (1 M, e.g., 50mL). The resulting homogeneous yellow solution is cooled to −25° C. andn-hexyllithium (4.3 eq, e.g., 202 mmol; 2.3 M in hexanes) is slowlyadded, after which the solution becomes dark red and is stirred at −20°C. for 20 min.

(S)-Tert-butyl 1-(methoxy(methyl)amino)-1-oxopropan-2-ylcarbamate (1.3eq, e.g., 61.0 mmol) is charged into a second dry round bottom flaskunder N₂ and suspended in 70 mL of dry THF and cooled to −10° C.Isopropyl magnesium chloride (2 M, 2.7 eq, e.g., 127 mmol) is slowlyadded resulting in a clear yellow solution. This solution is then slowlycanulated dropwise into the first round bottom flask. After addition iscomplete, the dark solution is slowly warmed to RT and stirred at RT for2 h. The reaction mixture is then recooled to −10° C. and quicklycanulated into another flask fitted with ethyl acetate (e.g., 15 mL) andisobutyric acid (e.g., 10 mL) at −10° C. under N₂. During this time themixture goes from orange and cloudy to clear and homogeneous. Afteraddition, the mixture is stirred for 5 min after which water (e.g., 10mL) is rapidly added and it is stirred vigorously for 10 min at RT.

The mixture is then transferred to a separation funnel, and water (e.g.,200 mL) is added to dissolve salts (pH˜9). The water layer is extractedwith EtOAc (e.g., 3×400 mL). The aqueous layer is then acidified withHCl (2 M) to pH 3, and then extracted with EtOAc (e.g., 3×500 mL), driedover sodium sulfate and concentrated to provide crude material which isfiltered under vacuum through a pad of silica gel using a MeOH/DCM(gradient of 2-10% MeOH) to provide the acid H-2 after concentration.

A 50 mL round bottom flask with a stir bar is filled with benzoic acidH-2 (1 eq., e.g., 14.63 mmol) in acetic anhydride (1.5 M, e.g., 10 mL)and then stirred at 70° C. for 2.5 hours until complete conversion tothe product is indicated by LC/MS. The acetic anhydride is evaporatedunder reduced pressure and the crude residue is purified with combiflash(gradient of EtOAc/hexanes) to give the lactone H-3.

A 50 mL dry round bottom flask with a stir bar is filled with amineR₂NH₂ (5.1 eq, e.g., 1.54 mmol) in 2 mL of DCM (0.8 M) after whichtrimethylaluminum (5.1 eq, e.g., 1.54 mmol) is added to the solution andstirred for 15 min. A solution of lactone H-3 (1.0 eq, e.g., 0.31 mmol)in DCM (1.5 M, e.g., 2 mL) is then added. The mixture is then stirred atRT for 3 h until LC/MS analysis showed complete formation of the desiredproduct. The reaction mixture is quenched with 10 mL of Rochelle's saltand stirred for 2 h. The mixture is then diluted with DCM, washed withbrine, dried with over sodium sulfate and evaporated to give a yellowsticky liquid H-4 which is used directly in next step.

To the amide H-4 (1 eq, e.g., 0.31 mmol) in isopropanol (0.06 M, e.g., 5mL) was added 3 mL of concentrated HCl (300 eq). The mixture is thenheated in an oil bath at 65° C. for 3 h until LC/MS shows no remainingstarting material. The flask is then removed from heat and the solventsare evaporated under reduced pressure to provide a yellow solid H-5which is used directly in subsequent transformations.

(iii) General Methods for Alkyne Synthesis:

Method I

A sealed vessled is chared with PdCl₂(MeCN)₂ and X-Phos (3:1 ratio ofX-Phos to PdCl₂(MeCN)₂, 5-15 mol % catalyst), cesium carbonate (1.5-3.0equiv) and propionitrile (0.5 M). The mixture is stirred for 5 min afterwhich the aryl bromide or aryl iodide substrate was added. After another5 minutes of stirring TMS-acetylene (3.0 equiv) is added and the flaskis sealed and heated at RT for 10 min followed by 1 h of heating at 95°C. The reaction is allowed to cool after which it is concentrateddirectly onto silica gel and purified using flash silica gelchromatography (gradient of ethyl acetate/hexanes) to provide alkyneI-1.

Alkyne I-1 (1.0 equiv) is then dissolved in tetrahydrofuran (0.13 M) andcharged with TBAF (1.1 equiv, 1.0 M in tetrahydrofuran). The resultingmixture is stirred at RT for 6 h after which it is poured into saturatedbicarbonate solution and extracted with ethyl acetate. The organic layeris washed with brine and concentrated onto silica gel where it ispurified directly by flash silica gel chromatography (gradient of ethylacetate/hexanes) to provide aryl alkyne 1-2.

Method J

Aldehyde (1.0 equiv) was a dissolved in anhydrous methanol (0.2-0.5 mM)and charged with cesium carbonate (1.0 equiv) and cooled to 0-5° C.Dimethyl (1-diazo-2-oxopropyl)phosphonate (1.0 equiv) was added dropwiseafter which the reaction was allowed to stir for 1-18 h after which thecrude mixture was concentrated onto silica gel and purified directly byflash silica gel chromatography to provide the desired alkyne J-1.

Method K

A secondary amine (1.0 equiv) is dissolved in acetonitrile (0.42 M) andpotassium carbonate (1.1 equiv) was added. The white suspension wasstirred at 0-5° C. for 5 min after which point propargyl bromide (1.01equiv) was added dropwise over 3 min. The reaction was then stirred foran additional 15 min at 0-5° C. and then at room temperature for 15 h.The heterogeneous mixture was then filtered. The filtrate wasconcentrated under reduced pressure, diluted with MTBE and washed withwater (2×), brine (1×), dried over sodium sulfate and then filteredthrough celite. The resulting filtrate was concentrated and purifiedusing flash silica gel chromatography to provide the desired alkyne K-1.

Example 1

Compound 4 was prepared in 3 steps from compound A according to thefollowing procedures: Compound A was prepared according to Method A. Itwas coupled to2-((tert-butoxycarbonyl)amino)pyrazolo[1,5-a]pyrimidine-3-carboxylicacid according to the following procedure: Compound A (27.4 mmol, 1.0equiv), HOBt hydrate (1.2 equiv),2-((tert-butoxycarbonyl)amino)pyrazolo[1,5-a]pyrimidine-3-carboxylicacid (1.05 equiv) and EDC (1.25 equiv) were added to a 200 mL roundbottomed flask with a stir bar. N,N-Dimethylformamide (50 mL) was addedand the suspension was stirred at RT for 2 min. Hunig's base (4.0 equiv)was added and after which the suspension became homogeneous and wasstirred for 22 h resulting in the formation of a solid cake in thereaction flask. The solid mixture was added to water (600 mL) andstirred for 3 h. The resulting cream colored solid was filtered andwashed with water (2×100 mL) and dried. The solid was then dissolved inmethylene chloride (40 mL) after which trifluoroacetic acid (10 equiv,20 mL) was added and the reaction was stirred for 30 min at RT afterwhich there is no more starting material by LC/MS analysis. The solutionwas then concentrated and coevaporated with a mixture of methylenechloride/ethanol (1:1 v/v) and then dried under high vacuum overnight.The resulting solid was triturated with 60 mL of ethanol for 1 h andthen collected via vacuum filtration. The beige solid was thenneutralized with sodium carbonate solution (100 mL) and then transferredto a separatory funnel with methylene chloride (350 mL). The water layerwas extracted with an additional 100 mL of methylene chloride. Thecombined organic layers were dried over sodium sulfate, filtered andconcentrated under vacuum to provide a pale yellow solid that waspurified using flash silica gel chromatography (Combiflash, 24 g column,gradient of 0-5% methanol/methylene chloride) to provide amide B. ESI-MSm/z: 459.4 [M+H]+.

Amide B was placed in a sealed tube (0.67 mmol, 1.0 equiv) followed bydichlorobis(acetonitrile)palladium (15 mol %), X-Phos (45 mol %), andcesium carbonate (3.0 equiv) Propionitrile (5 mL) was added and themixture was bubbled with Ar for 1 min. 4-Ethynyl-1-methyl-1H-pyrazole(1.24 equiv) was added and the resulting orange mixture was sealed andstirred in an oil bath at 85° C. for 1.5 h. The resulting brownish-blackmixture was allowed to cool at which point there was no more SM by LC/MSanalysis. The mixture was then filtered through a short plug of cottonusing acetonitrile and methylene chloride. The combined filtrates wereconcentrated onto silica gel and purified using flash silica gelchromatography (Combiflash, 4 g column, gradient of 0-5% methylenechloride/methanol). The resulting material was further purified byreverse phase HPLC (15-90% acetonitrile with 0.1% formic acid/water with0.1% formic water) to provide desired compound 4. ESI-MS m/z: 529.5[M+H]+.

The following compounds were prepared in analogous fashion. The alkyneswere either commercially available or prepared using Method I, J, or Kas described herein.

Compound no. Structure Alkyne ESI-MS m/z Compound 2

491.1 [M + H]⁺ Compound 5

525.5 [M + H]⁺ Compound 6

556.3 [M + H]⁺ Compound 7

529.5 [M + H]⁺ Compound 8

506.1 [M + H]⁺ Compound 9

489.4 [M + H]⁺ Compound 10

548.6 [M + H]⁺ Compound 11

507.1 [M + H]⁺ Compound 12

493.1 [M + H]⁺ Compound 13

479.1 [M + H]⁺ Compound 14

546.5 [M + H]⁺ Compound 15

493.4 [M + H]⁺ Compound 16

493.4 [M + H]⁺ Compound 17

526.5 [M + H]⁺ Compound 18

557.1 [M + H]⁺ Compound 19

543.2 Compound 20

556.2 [M + H]⁺ Compound 26

526.3 [M + H]⁺ Compound 28

556.3 [M + H]⁺ Compound 30

529.4 [M + H]⁺ Compound 32

526.4 [M + H]⁺ Compound 34

505.3 [M + H(−OEt)]⁺ Compound 35

543.4 [M + H]⁺ Compound 37

557.4 [M + H]⁺ Compound 38

543.4 [M + H]⁺ Compound 40

546.6 [M + H]⁺ Compound 41

532.6 [M + H]⁺ Compound 54

532.6 [M + H]⁺ Compound 56

561.7 [M + H]⁺ Compound 57

506.6 [M + H]⁺ Compound 59

532.5 [M + H]⁺ Compound 60

545.6 [M + H]⁺ Compound 61

540.3 [M + H]⁺ Compound 64

517.6 [M + H]⁺ Compound 65

531.6 [M + H]⁺ Compound 66

516.5 [M + H]⁺ Compound 67

540.3 [M + H]⁺ Compound 27

533.5[M + H]⁺ Compound 69

597.2 [M + H]⁺ Compound 73

529.2 2 [M + H]⁺ Compound 75

546.2 [M + H]⁺ Compound 76

546.2 [M + H]⁺ Compound 77

540.3 [M + H]⁺ Compound 78

546.2 [M + H]⁺ Compound 79

560.1 [M + H]⁺ Compound 81

529.0 [M + H]⁺ Compound 84

519.4 [M + H]⁺ Compound 85

546.5 [M + H]⁺ Compound 86

547.0 [M + H]⁺

Example 2

Compound A was prepared according to Method F. It was the converted tocompound AA1 using the analogous procedure for compound B in Example 1.Compound 1 was then prepared from compound AA1 in two steps according tothe following procedures: Compound AA1 (0.55 mmol, 1.0 equiv),PdCl₂(MeCN)₂ (10 mol %), X-Phos (30 mol %) and cesium carbonate (2.6equiv) were suspended in proprionitrile (4 mL). The mixture was bubbledwith Ar for 25 min after which trimethyl(propargyl)silane (1.3 equiv)was added and the reaction was sealed and heated to 90° C. The mixturewas allowed to heat for 4.5 h after which it was cooled and partitionedbetween ethyl acetate and water. The layers were separated and theaqueous layer was extracted with ethyl acetate (2×). The organic layerswere combined, dried over sodium sulfate and concentrated onto silicagel (2 g). The crude material was then purified using flash silica gelchromatography (ISCO Combiflash Si-12 g, gradient of 10-55%acetone/methylene chloride) to provide a mixture of compound B anddeprotected compound 1.

The mixture (0.23 mmol, 1.0 equiv) was redissolved in anhydroustetrahydrofuran (6 mL). TBAF in THF (1.0 M, 1.2 equiv) was added and theresulting mixture was stirred at RT for 45 min until complete conversionto compound 1 by TLC analysis. The reaction was then concentrated ontosilica gel (1 g) and purified by flash silica gel chromatography(Interchim Si-25 g HP silicycle, gradient of 14-45% acetone/methylenechloride) to provide compound 1. ESI-MS m/z: 464.1 [M+H]⁺.

Example 3

Compound kk was prepared from compound A (example 2) under standard Bocprotection conditions. It was then converted to compound 11 using theanalogous coupling procedure for compound B in Example 2 except that3,3-dimethylbut-1-yne was used in place of triethylsilylacetylene toprovide compound 11

Compound 11 (0.094 mmol, 1.0 equiv) was dissolved in anhydrous methylenechloride (2 mL). Trifluoroacetic acid (400 uL, 55 equiv) was added andthe reaction was allowed to stir at RT for 2 h until at which pointthere was no more SM by LC/MS analysis. The reaction was carefullyquenched with sodium bicarbonate solution and the aqueous layer wasextracted with methylene chloride (2×). The combined organic layers weredried with sodium sulfate and concentrated. The crude material waspurified using reverse phase chromatography (Interchim, gradient ofacetonitrile and water with 0.1% formic acid) to provide the free aminewhich was then coupled to2-((tert-butoxycarbonyl)amino)pyrazolo[1,5-a]pyrimidine-3-carboxylicacid using Method D followed by Boc-deprotection again using theanalogous conditions from Example 11 to provide the desired compound 3.ESI-MS m/z: 505.1 [M+H]⁺.

Example 4

A solution of 3-butyn-2-ol (10 mL, 128 mmol) in N,N-dimethylformamide(20 mL) was added over 30 minutes to a stirred slurry of sodium hydride(60% dispersion in mineral oil, (7.65 g, 2.5 equiv) inN,N-dimethylformamide (100 mL) at 0° C. under an argon atmosphere. After30 min, dimethyl sulfate (1.5 equiv) was added over 30 min at 0° C. Themixture was then stirred for 30 min at 0° C. after which acetic acid wasslowly added (1.05 equiv) and the reaction was allowed to warm to roomtemperature while stirring for an additional 2 h. The product wasisolated from fractional distillation directly from the reaction mixture(58-63° C.) to provided ether 4-a that was used directly in the nextstep. Compound 4-a was then coupled to compound A using analogousSonogashira conditions as in to Example 1 to generate compound 22.ESI-MS m/z: 507.5 [M+H]⁺.

Example 5

Compound 25 was prepared in analogous fashion to compound B inExample 1. It was then coupled to 4-ethynyl-1-methyl-1H-pyrazole usingthe Sonogashira conditions in Example 1 to provide compound 25. ESI-MSm/z: 493.4 [M+H]⁺.

Example 6

Compound 23 was prepared in analogous fashion to compound 25 in Example5 except that 5-ethynyl-1-methyl-1H-imidazole was used in place of4-ethynyl-1-methyl-1H-pyrazole. ESI-MS m/z: 493.4 [M+H]⁺.

Example 7

Compound 24 was prepared in analogous fashion to compound 25 in Example5 except that ethynylcyclopropane was used in place of4-ethynyl-1-methyl-1H-pyrazole. ESI-MS m/z: 453.4 [M+H]⁺.

Example 8

Compound 44 was isolated as a byproduct from Example 5. ESI-MS m/z:453.4 [M+H]⁺.

Example 9

Compound 21 was prepared from compound AA1 using analogous couplingconditions for the preparation of compound 4 in Example 1. ESI-MS m/z:530.2 [M+I-1]⁺.

Example 10

3-Aminopyrazine-2-carboxylic acid was coupled to compound A using MethodD to provide compound 10-a. It was then converted to compound 29 usinganalogous coupling conditions for the preparation of compound 4 inExample 1. ESI-MS m/z: 490.3 [M+H]⁺.

Example 11

Pyrazolo[1,5-a]pyrimidine-3-carboxylic acid was coupled to compound Ausing Method D to provide compound 11-a. It was then converted tocompound 39 using analogous coupling conditions for the preparation ofcompound 4 in Example 1. ESI-MS m/z: 514.4 [M+H]⁺.

Example 12

1,5-Naphthyridine-4-carboxylic acid was coupled to compound A usingMethod D to provide compound 12a. It was then converted to compound 42using analogous coupling conditions for the preparation of compound 4 inExample 1. ESI-MS m/z: 525.3 [M+H]⁺.

Example 13

Compound 13-a (0.058 mmol, 1.0 equiv) was dissolved in anhydrousacetonitrile (2 mL). Sodium iodide (1.5 equiv) was added followed byTMS-C1 (1.5 equiv) after which point the solution turned to a yellowsuspension. The mixture was then heated to 65° C. for 5 h after whichthere was no more starting material by LC/MS analysis. The reaction wasallowed to cool and poured into water (4 mL) and stirred for 15 minafter which it was partitioned between water and methylene chloride. Theorganic layer was when dried and concentrated. The crude material waspurified using reverse phase HPLC (Interchim, gradient of 10-90%acetonitrile/water with 0.1% formic acid) to provide desired compound31. ESI-MS m/z: 542.4 [M+H]⁺.

Example 14

Compound 33 was prepared from compound 14-a using the analogousconditions as in Example 13. ESI-MS m/z: 542.4 [M+H]⁺.

Example 15

Compound 34 was (0.47 mmol, 1.0 equiv) was dissolved in acetone (5 mL)and water (4 mL). p-Toluene sulfonic acid (25 mol %) was added and thecloudy mixture was heated to 50° C. The mixture was then allowed to coolafter which most of the solvent was removed under vacuum. The residuewas then partitioned between methylene chloride and saturated sodiumbicarbonate. The organic layer was separated and adsorbed onto SiO₂ (3g) after which it was purified by flash silica gel chromatography (ISCO,24 g Si column, gradient of 25-100% ethyl acetate/hexanes) to providethe desired aldehyde 36. ESI-MS m/z: 477.2 [M+H]⁺.

Example 16

5-Ethynyl-1H-pyrazole (1.1 mmol, 1.0 equiv) was dissolved in methylenechloride (10 mL). Triethylamine (3.0 equiv) and Boc anhydride (1.0equiv) were then added and the reaction was allowed to stir for 2 h.Water (100 mL) was added and the mixture was transferred to a separatoryfunnel. The layers were separated and the water layer was washed withwater (2×20 mL). The organic layers were dried over MgSO4 andconcentrated to provide alkyne 16-a which was used directly in the nextstep.

A pressure flask (15 mL) was charged with compound B (0.22 mmol, 1.0equiv), X-Phos (45 mol %), dichlorobis(acetonitrile)Pd (15 mol %), andcesium carbonate (1.1 equiv) under a flow of N₂. Propionitrile (3 mL)was added and the solution was bubble with Ar for 1 min. Alkyne 16-a(2.5 equiv) was then added followed by Boc anhydride (1.0 equiv) and thereaction was sealed and heated to 100° C. for 1 h. The reaction was thenfiltered and concentrated. The residue was redissolved in methylenechloride (3 mL) after which trifluoroacetic acid (800 uL) was added andthe mixture was stirred for 1 h. The reaction was then concentrated ontosilica gel and purified by flash silica gel chromatography (gradient0-30% methanol/methylene chloride) to provide compound 43. ESI-MS m/z:515.4 [M+H]⁺.

Example 17

Compound 17-a was prepared according to Method F. It was then convertedto Compound 55 in analogous fashion to compound 21 in Example 9. ESI-MSm/z: 468.3 [M+H]⁺.

Example 18

A sealed tube (30 mL) was charged with compound B (0.69 mmol, 1.0equiv), dichlorobis(acetonitrile)palladium (10 mol %), X-Phos (30 mol %)and cesium carbonate (1.5 equiv). Acetonitrile (10 mL) was addedfollowed by the additional of ethynyltrimethylsilane (0.4 mL) and themixture was purged with Ar for 1 min. The reaction was then sealted andheated in an oil bath to 85° C. After 45 min, an additional aliquot ofethynyltrimethylsilane (1.0 mL) was added and reheated to 75° C. for 14h after which there was no more starting material by LC/MS analysis. Themixture was filtered and concentrated onto silica gel and purified byflash silica gel chromatography (Combiflash, 12 g column, gradient of0-5% methanol/methylene chloride) to provide compound 18-a.

Compound 18-a (0.57 mmol, 1.0 equiv) was the dissolved intetrahydrofuran (4 mL). A solution of TBAF in tetrahydrofuran (0.8 mL,1.0 M) was added and the mixture was stirred at RT for 1 h at whichpoint the deprotected product was observed as the desired peak by LC/MSanalysis. The solution was concentrated onto silica gel and purifiedusing flash silica gel chromatography (Combiflash, 12 g column, gradientof 0-5% methanol/methylene chloride) to provide compound 18-b.

An oven dried RBF with a stir bar was charged with CuI (0.34 mmol, 1.0equiv), 1,10-phenanthroline (1.0 equiv) and KF (1.0 equiv). DryN,N-dimethylformamide (2 mL) was added and the mixture was stirred for15 min under an atmosphere of air. Trimethyl(trifluoromethyl)silane (5.0equiv) was then added and the mixture was heated to 100° C. under an airatmosphere. A solution of compound 18-b (1.0 equiv in 2 mLN,N-dimethylformamide) was added over the course of 4 h using a syringepump. Following the completion of compound 18-b addition, the reactionwas stirred for an additional 1.5 h at 100° C. At this point thereaction was allowed to cool after which water (100 mL) was added andthe mixture was extracted with methylene chloride (3×). The combinedorganics were washed with water, dried over sodium sulfate andconcentrated onto silica gel after which the material was purified byflash silica gel chromatography (Combiflash, 4 g column, gradient of0-10% methanol/methylene chloride). The crude material was furtherpurified by reverse phase HPLC (Interchim, gradient of 0-10%acetonitrile:water with 0.1% formic acid to provide the desired alkyne58. ESI-MS m/z: 517.5 [M+H]⁺.

Example 19

3-Quinuclidone hydrochloride (9.6 mmol, 1.0 equiv) was suspended inmethylene chloride (30 mL) and potassium carbonate solution was added(1.0 M, 16 ml). The mixture was stirred for 30 min after which theorganic later was collected and the aqueous layer was washed withmethylene chloride (3×20 mL), dried over sodium sulfate, filtered andconcentrated to provide the corresponding free base.

A solution of ethynyltrimethylsilane (10.6 mmol, 1.1 equiv) intetrahydrofuran (10 mL) was cooled to −10° C. n-Butyl lithium (2.5 M inTHF, 1.15 equiv) was added over 7 min. The reaction was stirred at −10°C. for 30 min after which it was cooled to −78° C. 3-Quinuclidone (1.0equiv in 20 mLTHF) was added to the flask over a period of 20 min,stirred for 15 additional min after which the cooling bath was removedand the reaction was allowed to stir at 23° C. for 15 h. The mixture wasthen quenced with saturated ammonium chloride (50 mL) and extracted withethyl acetate (5×25 mL). The combined organic layers were then washedwith water (1×20 mL) and brine (1×20 mL), dried over sodium sulfate andconcentrated under reduced pressure to provide alkyne 19-a which wasused directly in the next step.

Compound 19-a (7.7 mmol, 1.0 equiv) was dissolved in methanol (17 mL)and treated with potassium carbonate (1.05 equiv). The reaction wasallowed to stir at room temperature for 4 h after which it was filteredthrough celite, washing with 10% methanol in methylene chloride. Thefiltrates were concentrated under reduced pressure to half the volumeand filtered again after which they were concentrated completely underreduced pressure. The material was then redissolved in chloroform (30mL) and washed with 50% saturated brine (10 mL). The aqueous layer wasextracted with chloroform (3×20 mL). The combined organic layers werethen washed with brine (5 mL), dried over sodium sulfate andconcentrated under reduced pressure to provide compound 19-b.

An oven dried sealed tube was charged withdichlorobis(acetonitrie)palladium (15 mol %), X-Phos (45 mol %), andcesium carbonate (1.2 equiv) followed by propionitrile (5 mL). CompoundB (0.22 mmol, 1.0 equiv) was added and the reaction was degassed with Arfor 15 min. Alkyne 19-b (3.0 equiv) was added as a solid and the mixturewas purged for an additional 1 min with Ar. The flask was then sealedand heated to 100° C. for 2.5 h after which there was no more startingmaterial by LC/MS analysis. The mixture was filtered through celite andthe filtrate was concentrated under reduced pressure and adsorbed onto a1:4 ratio of Si-Triamine and silica gel (1.5 g) after which it waspurified using flash silica gel chromatography (Interchim, 12 g Sicolumn, gradient of 0-20% 1M ammonia in methanol/methylene chloride) toprovide the desired compound 62. ESI-MS m/z: 574.6 [M+H]⁺.

Example 20

Compound 20-a was prepared according to Method F. It was then coupled to2-((tert-butoxycarbonyl)amino)pyrazolo[1,5-a]pyrimidine-3-carboxylicacid according to Method D to provide compound 20-b. The Boc group wasdeprotected under standard conditions using trifluoroacetic acidaccording to the following procedure: Compound 20-b was dissolved in0.06 M methylene chloride. Trifluoroacetic acid (40 equiv) was thenadded and the reaction was allowed to stir at room temperature for 30min. The mixture was then poured into saturated sodium bicarbonatesolution and extracted with methylene chloride (2×). The combinedorganic layers were dried over Na₂SO₄, and concentrated to providecompound 20-c which was used directly in the next step.

A vial was then charged with compound 20-c (0.25 mmol, 1.0 equiv),cesium carbonate (3.0 equiv), PdCl₂(CH₃CN)₂ (30 mol %), X-Phos (15 mol%), propionitrile (3 mL) and DMSO (0.5 mL). The mixture was bubbled withArgon for 10 min after which TMS-acetylene (4.0 equiv) was added and thereaction was sealed and heated to 100° C. for 2 h until there was nomore starting material as indicated by LC/MS analysis. The reaction wasthen partitioned between ethyl acetate and brine. The water layer waswashed with ethyl acetate (1×). The combined organics were dried overNa₂SO₄ and concentrated to provide crude compound 20-d which was useddirectly in the next step.

Compound 20-d (0.25 mmol, 1.0 equiv) was dissolved in tetrahydrofuran(10 mL) after which 1M TBAF in tetrahydrofuran (4.0 equiv, 989 uL). Ater15 min there was no more SM by HPLC analysis. The crude reaction is thenpartioned between methylene chloride and water. The aqueous layer wasfirst extracted with methylene chloride (2×) and then diluted with 1NHCl and extracted with ethyl acetate (2×). All the organic layers weredried over Na₂SO₄ and concentrated to provide crude material which wasfirst purified by flash silica gel chromatography (Interchim Si-25 g HPsilicycle, gradient of 30-100 ethyl acetate/hexanes) to provide materialwhich was further purified by HPLC (30-90% methanol/0.1% trifluoroaceticacid in water) to provide compound 46. ESI-MS m/z: 450.3 [M+H]⁺.

Example 21

Compound 21-a was prepared according to Method F. It was then coupled toTES-acetylene according to the following procedure: A vial was thencharged with compound 21-a (0.48 mmol, 1.0 equiv), cesium carbonate (2.6equiv), PdCl₂(CH₃CN)₂ (10 mol %), X-Phos (30 mol %) and acetonitrile (2mL). The mixture was bubbled with Argon for 10 min after whichTES-acetylene (1.3 equiv) was added and the reaction was sealed andheated to 90° C. for 2 h until there was no more starting material asindicated by LC/MS analysis. The reaction was then partitioned betweenethyl acetate and brine. The water layer was washed with ethyl acetate(1×). The combined organics were dried over Na₂SO₄ and concentratedprovide crude compound 21-b which purified using flash silica gelchromatography (Interchim Si-25 g HP silicycle, gradient of 30-100 ethylacetate/hexanes).

Compound 21-b was then Boc-deprotected and coupled to3-amino-pyrazine-2-carboxylic acid using Method D to provide compound21-c. Compound 21-c (0.11 mmol, 1.0 equiv) was dissolved intetrahydrofuran (4 mL) and treated with 1M TBAF in tetrahydrofuran (3.0equiv, 320 uL). After 35 min there was no more starting material byLC/MS analysis. The crude mixture was concentrated, pre-adsorbed ontosilica gel and purified using flash silica gel chromatography (InterchimSi-12 g HP silicycle, gradient of 40-100 ethyl acetate/hexanes) toprovide compound 47 as the desired product. ESI-MS m/z: 411.3 [M+H]⁺.

Example 22

Compound 22-a was prepared according to Method F. A 2 dram vial was thencharged with compound 22-a (0.59 mmol, 1.0 equiv), cesium carbonate (2.6equiv), PdCl₂(CH₃CN)₂ (10 mol %), X-Phos (30 mol %) and propionitrile (2mL). The mixture was bubbled with Argon for 25 min after whichTES-acetylene (2.0 equiv) was added and the reaction was sealed andheated to 90° C. for 3 h until there was no more starting material asindicated by LC/MS analysis. The reaction was then partitioned betweenethyl acetate and brine. The water layer was washed with ethyl acetate(1×). The combined organics were dried over Na₂SO₄ and concentratedprovide crude compound 32 which purified using flash silica gelchromatography (Interchim Si-25 g HP silicycle, gradient of 0-30 ethylacetate/hexanes) to provide the desired material.

The TES group was removed and then Boc-deprotected provide amine 22-b.This was then coupled to2-((tert-butoxycarbonyl)amino)pyrazolo[1,5-a]pyrimidine-3-carboxylicacid using Method D followed by Boc-deprotection to provide the desiredcompound 48. ESI-MS m/z: 388.0 [M+H]⁺.

Example 23

Compound 23-a was prepared according to Method F. It was then convertedto amine. This was then coupled to2-((tert-butoxycarbonyl)amino)pyrazolo[1,5-a]pyrimidine-3-carboxylicacid using Method D followed by Boc-deprotection to provide the desiredcompound 50. ESI-MS m/z: 478.0 [M+H]⁺.

Example 24

Compound 23-b was coupled to 3-amino-pyrazine-2-carboxylic using MethodD to prepare compound 49. ESI-MS m/z: 429.0 [M+H]⁺.

Example 25

Compound 45 was prepared in analogous fashion as compound 49, using3,4-difluoroaniline instead of 4-fluoroaniline and using2-aminopyrazolo[1,5-a]pyrimidine-3-carboxylic acid instead of3-amino-pyrazine-2-carboxylic acid. ESI-MS m/z: 486.1 [M+H]⁺.

Example 26

Compound A was prepared according to WO 2008118468.

A mixture of chloride A (0.93 mmol, 1.0 equiv), phenylboronic acid (1.5equiv), Pd(PPh₃)₄

(5 mol %) and sodium carbonate (2 equiv) in dioxane/water (4/1 v/v, 65mL) was then degassed with Ar for 10 min. The resulting mixture washeated to 85° C. and stirred for 3 hr. The resulting suspension wascooled to RT, partitioned between ethyl acetate and a saturated aqueoussodium chloride solution. The organic phase was separated, dried withsodium sulfate, pre-adsorbed on silica gel and purified using silica gelchromatography with ethyl acetate and hexanes to afford compound B.ESI-MS m/z: 413.3 [M+H]⁺.

A mixture of phthalimide B (0.56 mmol, 1.0 equiv) and hydrazine (20equiv) in methanol (10 mL) was heated to 75° C. and stirred for 1 hr.The resulting mixture was concentrated, re-suspended in methylenechloride and filtered. The filtrate was concentrated to dryness toafford compound C. ESI-MS m/z: 283.3 [M+H]⁺.

Compound C (1.3 mmol, 1.0 equiv) was dissolved in N,N-dimethylformamide(5 mL) and charged with Hunig's base (2.0 equiv) and Boc anhydride (1.1equiv). The mixture was stirred at RT for 1 h after which there was nomore starting material by HPLC analysis. The reaction was then pouredinto brine and extracted with ethyl acetate. The organic layer as washedwith brine, dried over sodium sulfate and pre-adsorbed onto silica gel(2 g). The residue was then purified using flash silica gelchromatography (Interchim, Si-25 g, gradient of 10-30% ethylacetate/hexanes) to provide compound D. ESI-MS m/z: 383.1 [M+H]⁺.

Compound D (0.52 mmol, 1.0 equiv) was added to a 25 mL RBF containing asuspension of PdCl₂(MeCN)₂ (15 mol %), X-Phos (45 mol %) and cesiumcarbonate (3.0 equiv) in propionitrile (5 mL). The mixture was stirredfor 1 min after which TMS-propargylsilane (3.0 equiv) was added. Themixture was then stirred at RT for 30 min followed by heating to 95° C.for 1 h. LC/MS analysis showed conversion of the starting material toprimarily compound E after which the reaction was allowed to cool. Itwas then partitioned between ethyl acetate and water in a separatoryfunnel. The layers were separated and the aqueous layer was extractedwith ethyl acetate (1×). The combined organic layers were dried withsodium sulfate and pre-adsorbed onto silica gel (2 g). The resultingmaterial was purified using flash silica gel chromatography (ISCO, 25 gcolumn, gradient of 10-30% ethyl acetate/hexanes) to provide alkyne E.ESI-MS m/z: 387.1 [M+H]⁺.

Compound E was then Boc deprotected according to the followingprocedure: Compound E (0.19 mmol, 1.0 equiv) was dissolved in methylenechloride (4 mL) followed by the addition of trifluoroacetic acid (1 mL).The reaction was allowed to stir at RT for 90 min after which there wascomplete conversion of starting material by HPLC analysis. The reactionwas quenched with saturated sodium bicarbonate solution and extractedwith methylene chloride. The organic layer was concentrated over sodiumsulfate and concentrated. The resulting amine was then converted tocompound 51 using the analogous procedures for the conversion ofcompound A to B in Example 1. ESI-MS m/z: 447.1 [M+H]⁺.

Example 27

Compound 63 was prepared in analogous fashion to compound 4 in Example 1except that compound AA1 was used as starting material. ESI-MS m/z:547.2 [M+H]⁺.

Example 28

Compound 4 (0.12 mmol, 1.0 equiv) was dissolved in a mixture of ethanoland ethyl acetate (20 mL, 3:1 v/v). Palladium on carbon (19 mg, 10% Pd)was added and the reaction was placed under an atmosphere of H₂. Themixture was stirred at RT for 41 h after which it was filtered through afilter disk, concentrated and purified by flash silica gelchromatography (Combiflash, 4 g Si column, gradient of 0-5%methanol/methylene chloride) to provide alkene 53. ESI-MS m/z: 531.6[M+H]⁺.

Example 29

4-Ethynyl-1-methyl-1H-pyrazole (1.8 mmol, 1.0 equiv) and pinacolborane(5.0 equiv) were combined in toluene (8 mL) in a RBF under Ar.Carbonylchlorohydridotris(triphenylphosphine) ruthenium(II) (10 mol %)was added and the reaction was heated to 50° C. for 1.5 h after whichthere was no more starting material by LC/MS analysis. The solvent wasevaporated and the crude residue was transferred to a separatory funnelwith ethyl acetate (10 mL) and washed with saturated sodium bicarbonate(10 mL), water (10 mL) and brine (10 mL). The organic layer was driedover magnesium sulfate, concentrated and purified using flash silica gelchromatography (gradient 10-40% ethyl acetate/hexanes) to provide alkene29-a.

Compound B (0.22 mmol, 1.0 equiv), PdCl₂(Amphos)₂ (10 mol %) and sodiumcarbonate (2.0 equiv) were charged to a 4 mL vial under an Aratmosphere. A solution of compound 29-a in dioxane/water (1.5 equiv, 2mL solvent, 4:1 v/v) was added and the reaction was stirred at RT for 5min under Ar before heating to 85° C. for 1 h. The reaction was thenallowed to cool, diluted with methylene chloride (15 mL) and washed withwater (15 mL). The aqueous layer was then washed with additionalmethylene chloride (2×15 mL). The organic layers were combined and thenwashed with water (30 mL), brine (20 mL), dried over sodium sulfate andconcentrated to provide crude material which was first purified by flashsilica gel chromatography (Interchim Si-12 g, gradient of 0-5%methanol/methylene chloride) followed by purification using reversephase HPLC (Interchim C18-Sunfire column, acetonitrile/water/0.1% formicacid) to provide compound 52. ESI-MS m/z: 531.4 [M+H]⁺.

Example 30

Compound 68 is prepared according to the methods described herein.ESI-MS m/z: 504.2 [M+H]⁺.

Example 31

Compound B and trans-1-propen-1-ylboronic acid were coupled using theanalogous Suzuki coupling conditions in Example 29 to provide Compound70. ESI-MS m/z: 465.2 [M+H]⁺.

Example 32

Compound B and 4-ethynylpiperidine-1-carboxylic acid tert-butyl esterwere coupled using the Sonogashira coupling conditions in Example 1 toprovide compound 32a. Compound 32a was then dissolved in methylenechloride (0.007 M) followed by the addition of trifluoroacetic acid (10equiv). The reaction was allowed to stir for 2 h at RT after which itwas concentrated under vacuum. The residue was treated with excesssaturated sodium bicarbonate. The resulting residue was isolated viavacuum filtration and washed with excess water to provide Compound 72.ESI-MS m/z: 532.6 [M+H]⁺.

Example 33

Compound 74 was prepared in 3 steps according to the followingprocedures: tert-Butyl 3-formylazetidine-1-carboxylate was converted totert-butyl 3-ethynylazetidine-1-carboxylate according to Method J. Itwas then coupled to compound B and subsequently deprotected in analogousfashion to the synthesis of Compound 72 in Example 32. ESI-MS m/z: 504.5[M+H]⁺.

Example 34

Compound 80 was prepared in 4 steps from 1H-pyrazole-4-carbaldehydeaccording to the following procedures: 1H-pyrazole-4-carbaldehyde (2.1mmol, 1.0 equiv) was dissolved in 20 mL methylene chloride followed bythe addition of triethylamine (3.0 equiv) and trityl chloride (1.0equiv). The reaction was stirred at RT for 1 h after it was quenchedwith water (1 mL) and extracted with methylene chloride. The organiclayers were concentrated and purified using flash silica gelchromatography (gradient 0-30% methanol/methylene chloride with 0.5%triethylamine). 1-Trity-1H-pyrazole-4-carbaldehyde was then converted toit's corresponding alkyne using Method J after which it was coupled tocompound B using the analogous coupling conditions in Example 1. Theresulting compound was then deprotected under standard triflouroaceticacid in methylene chloride deprotection conditions after which it wasconcentrated and purified using flash silica gel chromatography (ISCO,gradient 0-5% methanol/methylene chloride with 0.05% triethylamine andthen repurified using reverse-phase HPLC (Interchim C18-Sunfire column,gradient of acetonitrile/water with 0.01% formic acid) to providecompound 80. ESI-MS m/z: 515.0 [M+H]⁺.

Example 35

Compound 82 was prepared in 3 steps according to the followingprocedures: N-Boc-4-ethynylpiperidine (3.8 mmol) was dissolved indioxane (10 mL) and HCl in dioxane (4M, 5.0 equiv) was added. Thereaction was allowed to stir at RT for 22 h. The mixture wasconcentrated under reduced pressure, diluted with 10 mL dioxane andreevaporated under reduced pressure. Diethylether (20 mL) was then addedand the mixture was revaporated to provide the HCl salt that was useddirectly in the next step. A suspension of the HCl salt (1.05 mmol, 1.0equiv) in methylene chloride (1 mL) was cooled to 0-5° C. in an icebath. Hunig's base (3.0 equiv) was added and then after a minute ofstirring acetic anhydride (2.0 equiv) was added. The mixture was allowedto stir for 1 h after which there was no more starting material by TLCanalysis. The reaction was then diluted with methylene chloride (5 mL),washed with 5% citric acid (1×2 mL), water (1×2 mL) dried over sodiumsulfate, and evaporated under reduced pressure. The crude residue waspurified using flash silica gel chromatography (ISCO, 4 g column, 0-50%ethyl acetate in methylene chloride) to provideN-acetyl-4-ethynylpiperidine which was coupled directly to compound Busing the analogous Sonogashira coupling conditions in example 1 toprovide compound 82. ESI-MS m/z: 574.5 [M+H]⁺.

Example 36

A suspension of 4-ethynyl piperidine HCl (1.1 mmol, 1.0 equiv) wassuspended in methylene chloride (1 mL) and cooled to 0-5° C. in an icebath. Hunig's base (3.0 equiv) was added and then after a minute ofstirring methanesulfonyl chloride (2.0 equiv) was added and the reactionwas allowed to stir for 1 h after which there was no more startingmaterial by LC/MS analysis. The mixture was then diluted with methylenechloride (5 mL) washed with 5% citric acid (1×2 mL), water (1×2 mL),dried over sodium sulfate and concentrated. The crude residue waspurified using flash silica gel chromatography (ISCO, 12 g Si column,gradient of 0-10% ethyl acetate/methylene chloride) to provideN-methanesulfonamide-4-ethynylpiperidine which was coupled directly tocompound B using the analogous Sonogashira coupling conditions inexample 1 to provide compound 83. ESI-MS m/z: 610.6 [M+H]⁺.

Example 37

Compound 88 was prepared in analogous fashion as compound 21 in example9 except that 4-ethynyl-1,5-dimethyl-1H-pyrazole was used in place of4-ethynyl-1-methyl-1H-pyrazole. A suspension of(S)-2-amino-N-(1-(5-chloro-4-oxo-3-phenyl-3,4-dihydroquinazolin-2-yl)ethyl)pyrazolo[1,5-a]pyrimidine-3-carboxamide(146 mg, 0.317 mmol), Cesium carbonate (198 mg, 0.608 mmol, 2 eq.),Dichlorobis(acetonitrile)palladium (II) (15 mg, 0.058 mmol, 0.2 eq.) andXphos (87 mg, 0.182, 0.6 eq.) in propionitrile (2 mL) was bubbled withargon for 5 minutes. The mixture was charged with4-ethynyl-1,5-dimethyl-1H-pyrazole (73 mg, 0.6 mmol, 2 eq.), heated to95° C. and stirred for 2 hr. The resulting mixture was cooled to RT,partitioned between Ethyl acetate and water. The organic phase wasseparated, washed with saturated aqueous sodium chloride solution, driedwith sodium sulfate and concentrated. The residue was purified withsilica gel chromatography using a gradient of DCM and MeOH to afford(S)-2-amino-N-(1-(5-((1,5-dimethyl-1H-pyrazol-4-yl)ethynyl)-4-oxo-3-phenyl-3,4-dihydroquinazolin-2-yl)ethyl)pyrazolo[1,5-a]pyrimidine-3-carboxamide.ESI-MS m/z: 544.2 [M+H]⁺.

Example 38

To a stirred mixture of 2-Methyl-1-naphthoic acid 1 (2.5 g, 13.4 mmols)and DMF (0.67 mL) in anhydrous chloroform was added thionyl chloride (1mL, 13.6 mmols) and the mixture was heated at reflux for 1 h. Thesolvents were evaporated, dissolved in 10 mL DCM and added to a biphasicmixture of aniline (2.5 mL, 27 mmols) in 40 mL DCM and 40 mL 1M aqueoussodium hydroxide solution. The mixture was stirred for 30 min, theaqueous layer was extracted with DCM (3×20 mL), washed with cold 1M HCl(20 mL), water (3×20 mL), brine (20 mL), dried and the solvents wereevaporated under reduced pressure and the crude solid (3.68 g, 92%) wasrecrystallized from DCM-hexanes to give 1.57 g of pure amide P2. M+H262.23; M−H 260.23. To a stirred mixture of amide P2 (1.05 g, 1 mmol, 1eq) in anhydrous THF (8 mL) at −10° C. under an argon atmosphere, asolution of hexyllithium in hexanes (3.93 mL, 9.04 mols, 2.25 eq) wasadded drop wise over 8 min while keeping the internal temperaturebetween −10° C. and −7° C. The resulting mixture is then stirred at −10°C. for 30 min.

To a stirred mixture of (S)-tert-butyl1-(methoxy(methyl)amino)-1-oxopropan-2-ylcarbamate (1.12 g, 4.82 mmols,1.2 eq) in anhydrous THF (8 mL) at −10° C. under an argon atmosphere, asolution of isopropylmagnesium chloride in THF (2.53 mL, 5.06 mmols,1.26 eq) was added drop wise over 7 min while keeping inner temperaturebetween −10° C. and −7° C. The resulting mixture was stirred at −10° C.for 30 min. This solution was then slowly added to above reactionmixture while keeping inner temperature between −10° C. and −13° C. Theresulting mixture is stirred at −10° C. for 1 h and then allowed to warmto room temp over a period of 1 h. The reaction mixture was added into abiphasic mixture of 20 mL 1M citric acid and 30 mL ethyl acetate at −5°C. to 0° C. The aqueous layer was extracted with ethyl acetate (3×20mL), washed with water and brine (20 mL), dried over sodium sulfate, thesolvents were removed in vacuo and the residue was purified bychromatography on silica gel (40 g, 0-50% EtOAc-Hexanes) to give 1.353 gof P3 as a solid. M+H 432.42; M−H 431.43.

A solution of 3 (1.1 g, 2.54 mmols) in 9 mL anisole was treated withtrifluoroacetic acid (1.52 mL, 20.3 mmols) and the mixture was heated at50 C for 18 h. The mixture was cooled, treated with 25 mL MTBE, theprecipitated solids were filtered, washed with MTBE (3×10 mL) and driedto give 1.07 g (2.5 mmols) the TFA salt of P4 as a solid.

200 mg of the TFA salt of 4 (0.467 mmols) was suspended in 6 mL DCM,treated with aqueous ammonium hydroxide solution (2 mL, ˜6%) for 30 min.The mixture was diluted with water (10 mL), extracted with DCM (2×5 mL),washed with water (5 mL), dried and the solvents were evaporated invacuo to give 149 mg (0.467 mmols) of crude P4. The crude P4 (120 mg,0.382 mmols), 2-Aminopyrazolo[1,5-a]pyrimidinecarboxylic acid (75 mg,0.42 mmols), HOBt (70 mg, 0.46 mmols), EDC (91 mg, 0.48 mmols) andHunig's base (0.27 mL, 1.53 mmols) in 3 mL DMF was stirred for 19 h. Themixture was slowly diluted with 6 mL methanol, heated to 50 C and cooledto room temperature. The precipitated solids was collected, washed withmethanol and dried to give 89 as a solid (154 mg). ESI-MS m/z: 475.46[M+H]⁺.

Example 39

Compound 90 was prepared according to amide formation methods generallyknown in the art. ESI-MS m/z: 548.31 [M+H]⁺.

Example 40

Compounds 91 and 92 were prepared.

Compound no. Structure ESI-MS m/z Compound 91

547.25 [M + H]⁺ Compound 92

531.31 [M + H]⁺

Example 41

Compounds 93-108 were prepared according to the procedure below.

A suspension of aryl chloride (0.03-0.06 mmol), cesium carbonate (1.2eq.), dichlorobis(acetonitrile)palladium (II) (0.05 eq.) and Xphos (0.15eq.) in acetonitrile (2 mL) was bubbled with argon for 5 minutes. Themixture was charged with 4-ethynyl-1-methyl-1H-pyrazole (2 eq.), heatedto 75° C. and stirred for 6 hr. The resulting mixture was cooled to RT,partitioned between ethyl acetate and water. The organic phase wasseparated, washed with saturated aqueous sodium chloride solution, driedwith sodium sulfate and concentrated. The residue was purified onsemi-prep HPLC (C-18) using a gradient of ACN/Water/Formic acid(9.9/90/0.1% to 49.9/50/0.1%) to afford the desired compound (confirmedby LCMS).

Compound ESI-MS m/z no Structure [M + H]⁺ 93

547.2 94

467.2 95

543.2 96

547.2 97

565.2 98

543.2 99

535.3 100

574.2 101

543.2 102

543.2 103

495.2 104

588.2 105

543.3 106

543.3 107

515.2 108

557.3 110

534.2

Example 42

In a MW compatible vial,(S)-3-(1-aminoethyl)-8-chloro-2-phenylisoquinolin-1(2H)-one (700 mg,2.343 mmol), (4-methoxyphenyl)methanamine (3.2 g, 23.4 mmol, 20 eq.) anddiisopropylethylamine (1.6 mL, 9.4 mmol, 4 eq.) were dissolved in NMP(12 mL). The vial was sealed and heated to 180° C. in a under MWirradiation and stirred for 6 hr. The reaction mixture was cooled to RT,partitioned between Ethyl acetate and water. The organic phase wasseparated, washed with saturated aqueous sodium chloride solution, driedwith sodium sulfate and concentrated. The residue was purified withsilica gel chromatography using a gradient of DCM and MeOH to afford(S)-3-(1-aminoethyl)-8-((4-methoxybenzyl)amino)-2-phenylisoquinolin-1(2H)-one.ESI-MS m/z: 400.1 [M+H]⁺.(S)-3-(1-aminoethyl)-8-((4-methoxybenzyl)amino)-2-phenylisoquinolin-1(2H)-one(720 mg, 1.8 mmol),2-((tert-butoxycarbonyl)amino)pyrazolo[1,5-a]pyrimidine-3-carboxylicacid (1.2 g, 4.31 mmol, 2.4 eq.), HOBt (700 mg, 4.57 mmol, 2.5 eq.) andEDC (800 mg, 4.17 mmol, 2.3 eq.) were suspended in DMF (30 mL). Thereaction mixture was charged with diisopropylethylamine (2 mL, 11.45mmol, 6.4 eq.) and stirred at RT for 1 hr. The reaction mixture waspartitioned between Ethyl acetate and water. The organic phase wasseparated, washed with saturated aqueous sodium chloride solution, driedwith sodium sulfate and concentrated. The residue was purified withsilica gel chromatography using a gradient of Ethyl acetate and hexanesand triturated with MeOH to afford (S)-tert-butyl(3-((1-(8-((4-methoxybenzyl)amino)-1-oxo-2-phenyl-1,2-dihydroisoquinolin-3-yl)ethyl)carbamoyl)pyrazolo[1,5-a]pyrimidin-2-yl)carbamate.ESI-MS m/z: 660.3 [M+H]⁺. (S)-tert-butyl(3-((1-(8-((4-methoxybenzyl)amino)-1-oxo-2-phenyl-1,2-dihydroisoquinolin-3-yl)ethyl)carbamoyl)pyrazolo[1,5-a]pyrimidin-2-yl)carbamate(360 mg, 0.546 mmol) and anisole (238 μL, 2.183 mmol, 4 eq.) wasdissolved TFA (2 mL) and stirred at 60° C. for 1 hr. The reactionmixture was poured in a saturated aqueous bicarbonate solution. Theorganic phase was dried with sodium sulfate and concentrated. Theresidue was purified with silica gel chromatography using a gradient ofDCM. The residue was purified on semi-prep HPLC (C-18) using a gradientof ACN/Water/Formic acid to afford(S)-2-amino-N-(1-(8-amino-1-oxo-2-phenyl-1,2-dihydroisoquinolin-3-yl)ethyl)pyrazolo[1,5-a]pyrimidine-3-carboxamide.ESI-MS m/z: 440.2 [M+H]⁺.

Biological Activity Assessment

TABLE 15 In Vitro IC₅₀ data for selected compounds. PI3K δ/ RAJI δ/ RAJIRaw264.7 PI3K γ Raw264.7 Compound PI3K PI3K PI3K PI3K p110 δ p110 γ IC₅₀γ IC₅₀ no. α IC₅₀ β IC₅₀ δ IC₅₀ γ IC₅₀ assay IC₅₀ assay IC₅₀(selectivity) (selectivity) 1 D2 C2 B2 A3 A4 A5 X X 2 D2 D2 D2 C3 C4 A5X Y 3 D2 D2 D2 D3 D4 B5 W X 4 C2 C2 D2 A3 B4 A5 Y Y 5 D2 D2 A2 D3 A4 A5V W 6 D2 D2 D2 B3 C4 A5 Y X 7 D2 D2 D2 B3 C4 A5 Y Y 8 D2 D2 D2 C3 D4 C5X W 9 C2 C2 C2 B3 B4 A5 X Y 10 D2 D2 D2 B3 D4 A5 X X 11 D2 D2 D2 B3 D4B5 X X 12 D2 C2 C2 A3 B4 A5 X X 13 D2 C2 B2 A3 A4 A5 X W 14 C2 C2 A2 A3A4 A5 X W 15 D2 D2 B2 A3 B4 A5 X W 16 D2 C2 C2 A3 B4 A5 Y X 17 D2 D2 D2B3 B4 A5 Y Y 18 D2 D2 D2 B3 B4 A5 Y X 19 D2 D2 D2 B3 C4 A5 Y Y 20 D2 D2C2 A3 B4 A5 X X 21 D2 D2 D2 A3 B4 A5 Y Y 22 D2 D2 D2 B3 D4 A5 X X 23 C2C2 D2 D3 B4 A5 W W 24 C2 C2 C2 D3 B4 A5 W W 25 C2 C2 D2 C3 B4 A5 X X 26D2 D2 D2 B3 B4 A5 X Y 27 D2 D2 D2 B3 A4 A5 X Y 28 D2 D2 D2 D3 B4 A5 W X29 D2 C2 C2 B3 A4 A5 X W 30 D2 D2 D2 B3 B4 A5 Y Y 31 D2 D2 D2 B3 B4 B5 XW 32 D2 D2 D2 B3 C4 A5 Y Y 33 D2 D2 D2 A3 B4 A5 Y W 34 D2 D2 D2 C3 C4 B5X X 35 D2 D2 D2 B3 C4 A5 Y Y 36 C2 A2 C2 A3 B4 C5 X W 37 D2 D2 D2 D3 D4A5 W Y 38 D2 D2 D2 A3 C4 A5 Y Y 39 D2 D2 D2 B3 D4 B5 X X 40 C2 D2 D2 A3B4 A5 Y Y 41 D2 D2 D2 B3 B4 A5 Y Y 42 D2 D2 D2 B3 C4 A5 X X 43 D2 D2 D2B3 B4 A5 Y X 44 D2 C2 C2 D3 A4 B5 W V 45 D2 D2 D2 B3 B4 A5 Y W 46 D2 C2C2 A3 A4 A5 Y W 47 C2 A2 A2 A3 ND ND V ND 48 C2 B2 C2 C3 A4 A5 W V 49 D2C2 C2 A3 A4 A5 X V 50 D2 D2 C2 A3 A4 A5 Y W 51 D2 C2 C2 B3 A4 A5 W W 52D2 D2 D2 B3 C4 A5 Y Y 53 D2 D2 D2 D3 C4 B5 W X 54 C2 C2 C2 A3 B4 A5 Y W55 C2 C2 D2 D3 C4 A5 W X 56 D2 D2 D2 B3 B4 C5 X V 57 D2 D2 D2 C3 B4 B5 WW 58 D2 D2 D2 D3 C4 C5 W V 59 D2 D2 D2 B3 B4 A5 X X 60 D2 D2 D2 B3 B4 A5X Y 61 D2 D2 D2 C3 D4 A5 X Y 62 D2 D2 D2 C3 D4 C5 X V 63 D2 D2 D2 A3 C4A5 Y Y 64 D2 D2 D2 D3 D4 C5 W X 65 D2 D2 D2 D3 D4 C5 W X 66 D2 C2 C2 A3B4 A5 X X 67 D2 D2 D2 D3 D4 C5 W X 68 D2 D2 D2 D3 D4 C5 W W 69 D2 D2 D2D3 ND C5 W ND 70 D2 D2 D2 B3 A4 A5 X X 71 D2 D2 D2 E3 D4 ND V ND 72 D2D2 D2 C3 D4 C5 X W 73 D2 D2 D2 B3 C4 A5 X Y 74 D2 D2 C2 C3 D4 C5 W W 75D2 D2 D2 D3 D4 A5 W Y 76 D2 D2 D2 B3 B4 A5 Y X 77 D2 D2 D2 A3 C4 A5 Y Y78 D2 D2 D2 B3 B4 A5 X X 79 D2 D2 D2 C3 D4 A5 X Y 80 C2 C2 D2 A3 B4 A5 YY 81 C2 C2 C2 A3 ND ND Y ND 82 D2 D2 D2 B3 C4 A5 X X 83 D2 D2 D2 C3 C4A5 X X 84 D2 D2 D2 A3 B4 A5 Y X 85 D2 D2 D2 C3 ND ND X ND 86 D2 C2 C2 B3ND ND X ND 87 D2 D2 D2 E3 ND ND V ND 88 D2 D2 D2 B3 B4 A5 X Y 89 D2 D2D2 C3 D4 A5 X Y 90 D2 D2 D2 D3 D4 C5 W X 91 D2 D2 D2 C3 B4 C5 W W 92 D2D2 C2 C3 B4 C5 W V 93 D2 D2 D2 A3 ND ND Y ND 94 C2 B2 D2 B3 ND ND X ND95 D2 D2 D2 B3 ND ND X ND 96 C2 D2 D2 A3 ND ND Y ND 97 D2 D2 D2 B3 ND NDX ND 98 D2 D2 D2 B3 ND ND X ND 99 D2 D2 D2 D3 ND ND X ND 100 C2 C2 D2 A3ND ND Y ND 101 D2 D2 D2 A3 ND ND Y ND 102 D2 D2 D2 B3 ND ND X ND 103 D2D2 D2 C3 ND ND X ND 104 C2 C2 D2 A3 ND ND Y ND 105 C2 D2 D2 A3 ND ND YND 106 D2 C2 D2 A3 ND ND Y ND 107 D2 D2 D2 D3 ND ND X ND 108 D2 D2 D2 B3ND ND X ND 109 D2 C2 C2 A3 ND ND X ND 110 D2 D2 D2 A3 ND ND Y NDThe data in Table 15 are coded as follows.

For PI3K α, β, and δ RAJI p110 δ assay Raw264.7 p110 γ IC₅₀: For PI3K γIC₅₀: IC₅₀ assay IC₅₀ A2 = 1 to <500 nM A3 = 1 to <100 nM A4 = 1 to <100nM A5 = 1 to <50 nM B2 = 500 to <1000 nM B3 = 100 to <500 B4 = 100 to<500 nM B5 = 50 to <100 nM nM C2 = 1000 to <5000 C3 = 500 to <1000 C4 =500 to <1000 nM C5 = 100 to <10000 nM nM nM D2 = 5000 to 10000 D3 = 1000to 5000 D4 = 1000 to 10000 nM nM nM E3 = >5000 nM δ/γ IC₅₀ selectivity:ND = not determined V = 0.1 to 1 W = >1 to <10 X = 10 to <50 Y = 50 to<850

Example 222: PI3-Kinase HTRF™ Assay

A PI3-Kinase HTRF® assay kit (cat No. 33-016) purchased from MilliporeCorporation was used to screen compounds provided herein. This assayused specific, high affinity binding of the GRP1 pleckstrin homology(PH) domain to PIP3, the product of a Class 1A or 1B PI3 Kinase actingon its physiological substrate PIP2. During the detection phase of theassay, a complex was generated between the GST-tagged PH domain andbiotinylated short chain PIP3. The biotinylated PIP3 and the GST-taggedPH domain recruited fluorophores (Streptavidin-Allophycocyanin andEuropium-labeled anti-GST respectively) to form the fluorescenceresonance energy transfer (FRET) architecture, generating a stabletime-resolved FRET signal. The FRET complex was disrupted in acompetitive manner by non-biotinylated PIP3, a product formed in the PI3Kinase assay.

PI3 Kinase α, β, γ or δ activity was assayed using the PI3 Kinase HTRF®assay kit (catalogue No. 33-016) purchased from Millipore Corporation.Purified recombinant PI3Kα (catalogue No. 14-602-K), PI3Kβ (catalogueNo. 14-603-K), PI3Kγ (catalogue No. 14-558-K), and PI3Kδ (catalogue No.14-604-K) were obtained from Millipore Corporation. Purified recombinantPI3K enzyme was used to catalyze the phosphorylation ofphosphatidylinositol 4,5-bisphosphate (PIP2 at 10 μM) tophosphatidylinositol 3,4,5-trisphosphate (PIP3) in the presence of 10 μMATP. The assay was carried out in 384-well format and detected using aPerkin Elmer EnVision Xcite Multilabel Reader. Emission ratios wereconverted into percent inhibitions and imported into GraphPad Prismsoftware. The concentration necessary to achieve inhibition of enzymeactivity by 50% (IC₅₀) was calculated using concentrations ranging from20 μM to 0.1 nM (12-point curve). IC₅₀ values were determined using anonlinear regression model available in GraphPad Prism 5.

Example 223: Chemical Stability

The chemical stability of one or more subject compounds is determinedaccording to standard procedures known in the art. The following detailsan exemplary procedure for ascertaining chemical stability of a subjectcompound. The default buffer used for the chemical stability assay isphosphate-buffered saline (PBS) at pH 7.4; other suitable buffers can beused. A subject compound is added from a 100 μM stock solution to analiquot of PBS (in duplicate) to give a final assay volume of 400 μL,containing 5 μM test compound and 1% DMSO (for half-life determination atotal sample volume of 700 μL is prepared). Reactions are incubated,with shaking, for 24 hours at 37° C.; for half-life determinationsamples are incubated for 0, 2, 4, 6, and 24 hours. Reactions arestopped by adding immediately 100 μL of the incubation mixture to 100 μLof acetonitrile and vortexing for 5 minutes. The samples are then storedat −20° C. until analysis by HPLC-MS/MS. Where desired, a controlcompound or a reference compound such as chlorambucil (5 μM) is testedsimultaneously with a subject compound of interest, as this compound islargely hydrolyzed over the course of 24 hours. Samples are analyzed via(RP)HPLC-MS/MS using selected reaction monitoring (SRM). The HPLCconditions consist of a binary LC pump with autosampler, a mixed-mode,C12, 2×20 mm column, and a gradient program. Peak areas corresponding tothe analytes are recorded by HPLC-MS/MS. The ratio of the parentcompound remaining after 24 hours relative to the amount remaining attime zero, expressed as percent, is reported as chemical stability. Incase of half-life determination, the half-life is estimated from theslope of the initial linear range of the logarithmic curve of compoundremaining (%) vs. time, assuming first order kinetics.

Example 224: Expression and Inhibition Assays of p110α/p85α, p110β/p85α,p110δ/p85α, and p110γ

Class I PI3-Ks can be either purchased (p110α/p85α, p110β/p85α,p110δ/p85α from Upstate, and p110γ from Sigma) or expressed aspreviously described (Knight et al., 2004). IC₅₀ values are measuredusing either a standard TLC assay for lipid kinase activity (describedbelow) or a high-throughput membrane capture assay. Kinase reactions areperformed by preparing a reaction mixture containing kinase, inhibitor(2% DMSO final concentration), buffer (25 mM HEPES, pH 7.4, 10 mMMgCl₂), and freshly sonicated phosphatidylinositol (100 μg/ml).Reactions are initiated by the addition of ATP containing 10 μCi ofγ-32P-ATP to a final concentration of 10 or 100 μM and allowed toproceed for 5 minutes at room temperature. For TLC analysis, reactionsare then terminated by the addition of 105 μL 1N HCl followed by 160 μLCHCl₃:MeOH (1:1). The biphasic mixture is vortexed, briefly centrifuged,and the organic phase is transferred to a new tube using a gel loadingpipette tip precoated with CHCl₃. This extract is spotted on TLC platesand developed for 3-4 hours in a 65:35 solution of n-propanol:1M aceticacid. The TLC plates are then dried, exposed to a phosphorimager screen(Storm, Amersham), and quantitated. For each compound, kinase activityis measured at 10-12 inhibitor concentrations representing two-folddilutions from the highest concentration tested (typically, 200 μM). Forcompounds showing significant activity, IC₅₀ determinations are repeatedtwo to four times, and the reported value is the average of theseindependent measurements.

Other commercial kits or systems for assaying PI3-K activities areavailable. The commercially available kits or systems can be used toscreen for inhibitors and/or agonists of PI3-Ks including, but notlimited to, PI 3-Kinase α, ρ, δ, and γ. An exemplary system is PI3-Kinase (human) HTRF™ Assay from Upstate. The assay can be carried outaccording to the procedures suggested by the manufacturer. Briefly, theassay is a time resolved FRET assay that indirectly measures PIP3product formed by the activity of a PI3-K. The kinase reaction isperformed in a microtiter plate (e.g., a 384 well microtiter plate). Thetotal reaction volume is approximately 20 μL per well. In the firststep, each well receives 2 μL of test compound in 20% dimethylsulphoxideresulting in a 2% DMSO final concentration. Next, approximately 14.5 μLof a kinase/PIP2 mixture (diluted in 1× reaction buffer) is added perwell for a final concentration of 0.25-0.3 μg/mL kinase and 10 μM PIP2.The plate is sealed and incubated for 15 minutes at room temperature. Tostart the reaction, 3.5 μL of ATP (diluted in 1× reaction buffer) isadded per well for a final concentration of 10 μM ATP. The plate issealed and incubated for 1 hour at room temperature. The reaction isstopped by adding 5 μL of Stop Solution per well and then 5 μL ofDetection Mix is added per well. The plate is sealed, incubated for 1hour at room temperature, and then read on an appropriate plate reader.Data is analyzed and IC₅₀s are generated using GraphPad Prism 5.

Example 225: B Cell Activation and Proliferation Assay

The ability of one or more subject compounds to inhibit B cellactivation and proliferation is determined according to standardprocedures known in the art. For example, an in vitro cellularproliferation assay is established that measures the metabolic activityof live cells. The assay is performed in a 96 well microtiter plateusing Alamar Blue reduction. Balb/c splenic B cells are purified over aFicoll-Paque™ PLUS gradient followed by magnetic cell separation using aMACS B cell Isolation Kit (Miletenyi). Cells are plated in 90 μL at50,000 cells/well in B Cell Media (RPMI+10% FBS+Penn/Strep+50 μM bME+5mM HEPES). A compound provided herein is diluted in B Cell Media andadded in a 10 μL volume. Plates are incubated for 30 min at 37° C. and5% CO₂ (0.2% DMSO final concentration). A 50 μL B cell stimulationcocktail is then added containing either 10 μg/mL LPS or 5 μg/mL F(ab′)2Donkey anti-mouse IgM plus 2 ng/mL recombinant mouse IL4 in B CellMedia. Plates are incubated for 72 hours at 37° C. and 5% CO₂. A volumeof 15 μL of Alamar Blue reagent is added to each well and plates areincubated for 5 hours at 37° C. and 5% CO₂. Alamar Blue fluoresce isread at 560Ex/590Em, and IC₅₀ or EC₅₀ values are calculated usingGraphPad Prism 5.

Example 226: Tumor Cell Line Proliferation Assay

The ability of one or more subject compounds to inhibit tumor cell lineproliferation can be determined according to standard procedures knownin the art. For instance, an in vitro cellular proliferation assay canbe performed to measure the metabolic activity of live cells. The assayis performed in a 96-well microtiter plate using Alamar Blue reduction.Human tumor cell lines are obtained from ATCC (e.g., MCF7, U-87 MG,MDA-MB-468, PC-3), grown to confluency in T75 flasks, trypsinized with0.25% trypsin, washed one time with Tumor Cell Media (DMEM+10% FBS), andplated in 90 μL at 5,000 cells/well in Tumor Cell Media. A compoundprovided herein is diluted in Tumor Cell Media and added in a 10 μLvolume. Plates are incubated for 72 hours at 37° C. and 5% CO₂. A volumeof 10 μL of Alamar Blue reagent is added to each well and plates areincubated for 3 hours at 37° C. and 5% CO₂. Alamar Blue fluoresce isread at 560Ex/590Em, and IC₅₀ values are calculated using GraphPad Prism5.

Example 227: Antitumor Activity In Vivo

The compounds described herein can be evaluated in a panel of human andmurine tumor models. In one aspect, compounds provided herein may beevaluated in the following models according to methods known in the art.The dosage and schedule of administration may be varied depending on themodel. The results may be evaluated with those of selective deltainhibitors, and combinations of delta and gamma inhibitors, and/or withantibodies that block specific inhibitory receptors.

Paclitaxel-Refractory Tumor Models

1. Clinically-Derived Ovarian Carcinoma Model.

This tumor model is established from a tumor biopsy of an ovarian cancerpatient. Tumor biopsy is taken from the patient. The compounds describedherein are administered to nude mice bearing staged tumors using anevery 2 days×5 schedule.

2. A2780Tax Human Ovarian Carcinoma Xenograft (Mutated Tubulin).

A2780Tax is a paclitaxel-resistant human ovarian carcinoma model. It isderived from the sensitive parent A2780 line by co-incubation of cellswith paclitaxel and verapamil, an MDR-reversal agent. Its resistancemechanism has been shown to be non-MDR related and is attributed to amutation in the gene encoding the beta-tubulin protein. The compoundsdescribed herein can be administered to mice bearing staged tumors on anevery 2 days×5 schedule.

3. HCT116/VM46 Human Colon Carcinoma Xenograft (Multi-Drug Resistant).

HCT116/VM46 is an MDR-resistant colon carcinoma developed from thesensitive HCT116 parent line. In vivo, grown in nude mice, HCT116/VM46has consistently demonstrated high resistance to paclitaxel. Thecompounds described herein can be administered to mice bearing stagedtumors on an every 2 days×5 schedule.

One or more compounds as provided herein can be used in combination withother therapeutic agents in vivo in the multidrug resistant human coloncarcinoma xenografts HCT/VM46 or any other model known in the artincluding those described herein.

4. M5076 Murine Sarcoma Model

M5076 is a mouse fibrosarcoma that is inherently refractory topaclitaxel in vivo. The compounds described herein can be administeredto mice bearing staged tumors on an every 2 days×5 schedule.

Pancreatic Models

KPC model is a transgenic mouse model of pancreatic ductaladenocarcinoma (PDA), in which there is conditional expression of bothmutant KrasG12D and p53R172H alleles in pancreatic cells. Tumors developspontaneously in this mouse over a period of 3-6 months, and can be usedto study prophylactic, as well as therapeutic efficacy with novelagents. Cells from these KPC tumors can also be adoptively transferredinto syngeneic B6.129 hybrid mice, creating a model with a shorterlatency period and allowing large number of animals with tumors to besynchronously established. See e.g., Cancer Cell 7:468 (2005). In someembodiments, compounds provided herein can be administered alone or incombination with anti-PD-L1.

Pan02 model: The murine pancreatic adenocarcinoma cell line Pan02 is anonmetastatic tumor line, syngeneic to C57BL/6. It can be studiedfollowing s.c. injection into flank, or orthotopically followinginjection directly into the pancreas. See e.g., Cancer Res. 44: 717-726(1984).

Lung Models

LLC Lewis Lung carcinoma model: LLC cells are derived from a spontaneouslung tumor from a C57BL/6 mouse and can be studied as a s.c. tumor wheninjected in the flank, or as an orthotopic tumor if injected i.v.,following which it localizes to the lung.

LLC cells have also been modified to express a peptide from ovalbumin(LL2-OVA cells). Use of these cells, following either s.c. or i.v.injection, allows the tracking of OVA-specific CD8+ lymphocyctes andmeasurement of effects of therapy on the adaptive immune responseagainst the tumor. See e.g., Science 330:827 (2010).

Breast Model

The 4T1 mammary carcinoma is a transplantable tumor cell line that growsin syngeneic BALB/c mice. It is highly tumorigenic and invasive and,unlike most tumor models, can spontaneously metastasize from the primarytumor in the mammary gland to multiple distant sites including lymphnodes, blood, liver, lung, brain, and bone. See e.g., Current Protocolsin Immunology Unit 20.2 (2000). The parental 4T1 cell line has beenmodified to constitutively express luciferase (4T1-luc). 4T1-Luc cellline can be introduced orthotopically into the mammary fat pad of Balb/cmice. When introduced orthotopically the 4T-luc model grows rapidly atthe primary site and forms metastases at distal sites over a period of3-6 weeks. The rapid and efficient metastasis to organs affected inhuman breast cancer makes this an excellent mouse model for the study ofmetastatic progression of breast cancer in humans. Because the model issyngeneic to Balb/c mice, it can be used to study the role of the immunesystem in tumor growth and metastasis. See e.g., Cancer Res. 1992 Mar.15; 52(6):1399-405. For example, the cancer cells of the 4T1 model canmetastasize to the lung. 4T1 tumor cells are 6-thioguanine-resistant,and the metastatic cells can be detected and quantified by explantingorgans, plating dissociated cells in medium supplemented with6-thioguanine, and counting the number of 6-TG-resistant clonogenictumor cells. The 4T1 model can be a model of lung metastasis. See e.g.,Current Protocols in Immunology Unit 20.2 (2000); and Kerbel, R. S., TheBreast 22 (2013), S57-S65.

4T1-Luc, luciferin-expressing 4T1 cell line is cultured in filteredRPMI-1640 supplemented with 10% heat inactivated fetal bovine serum at37° C. in 5% CO₂ atmospheric air. 5000 4T1-Luc cells in 50 uL phosphatebuffered saline are implanted orthotopically into the mammary fat pad of6-8 week old Balb/c female mice. When tumors reach approximately 50-100mm³, mice are randomized into treatment groups. Mice are orallyadministered vehicle or an exemplary PI3Kγ inhibitor disclosed hereinthat has a delta/gamma selectivity ratio of greater than about 50. Forexample, the PI3Kγ inhibitor is orally administered 3 mg/kg once dailyfor 21 consecutive days alone or in combination with anti-PD-L1 (200 ug)or Isotype control (Rat IgG2b κ; 200 ug) antibodies that areadministered intraperitonal once every 3 days for a total of 5 doses.Tumor and body weight measurements are taken 3 times a week. Luciferinmeasurements will be taken twice a week imaged using the IVIS 200. Atthe end of the study, mice are euthanized and tumor will be harvestedfor evaluating pathway inhibition and immune response of constant PI3Kγpathway suppression. Plasma is collected for pharmokinetic (PK)analysis. Efficacy is determined, for example, by tumor volumemeasurements.

Another breast model is described as follows. Prior to injection, PyMT8119 cells are cultured in F12K supplemented with 5% Fetal Clone II, 50ug/ml gentamycin, 2.5 ug/ml fugizone, and 1 ul/ml MITO. PyMT 8119 cellsare injected subcutaneously or orthotopically in to the mammary fatpad(1×10{circumflex over ( )}6 cells/mouse) into C57B16 or nude mice. Tumormeasurements are recorded 3 times a week starting at d7 post injectionand tumor volumes are calculated using the formula:(length×width×width)/2=volume where length denotes the longest dimensionmeasured and width denotes the shortest dimension.

Lymphoma Model

EL4 is a C57BL/6 T thymoma and EG7 is an OVA-expressing subclone of EL4.The parental EL4 line has been modified to constitutively expressluciferase, which allows non-invasive imaging of tumor growth throughoutthe animal using the Xenogen imaging platform.

A20 is a Balb/c B cell lymphoma cell line derived from a spontaneousneoplasm found in an old Balb/cAnN mouse, expressing MHC class I andclass II H-2d molecules. The parental A20 cell line has been modified toconstitutively express luciferase(A20-Luc). A20-Luc, aluciferase-expressing cell line is derived from the parental A20 cellline can be implanted into Balb/c mice subcutaneously or intravenously.The systemic i.v. syngeneic model can be used to study the role of theimmune system in tumor growth and metastasis. See e.g., J. Clin Invest.2013; 123(6):2447-2463.

Melanoma Model

B16 murine melanoma cells are syngeneic with C57BL/6 mice and can bestudied after s.c., i.d. (inter dermal), or i.v. injection. Placement ateither site will result in metastases to lung and other organs. Thismodel has been extensively studied in terms of the role that inhibitoryreceptors play in the anti-tumor immune response. See e.g., PNAS107:4275 (2010).

The B16-F10 luc tagged tumor model allows evaluation of efficacy andpharmacodynamic effects of PI3K-γ inhibitors and other anti-tumoragents. The model can be used as follows. C57 BLK mice (male) areordered at 5-6 weeks from Jackson Labs, n=40. Mice are dosed QD with aPI3K-γ inhibitor. PDL-1 dose E3D or vehicle is dosed QD. Tumormeasurements and bodyweights are taken 3×/week. Tumors are harvested onthe last day or if the vehicle reached approximately 2000 mm³. Tumorsare cut in half. One half is cut in half again with ½ fixed in 10% NBFthe other frozen in OCT for frozen sections. The remaining half isprocessed to single cell suspention and evaluated by FACS using twopanels. Dosing is listed in Table 16 below. Vaccination of this modelcan enhance response to treatment. See e.g., Duraiswamy, J. et al.,Cancer Res, 73(12), 2013, 3591; Curran, M. A., et al., PNAS, 107(9),2010, 4275.

TABLE 16 Dose Drug vol conc Dose (ml/ (mg/ Grp # N Compound (mg/kg)Route Frequency kg) ml) 1 12 Vehicle  0 PO/IP QD 5/10 0 2 12 Compound 415 PO QD  5 3 4 12 PDL-1 200 uL/ IP Q3D 10 — mouse

Melanoma is known to be sensitive to immunotherapies, and data linking apoor prognosis to high TAM cell counts in these tumors have beenreported. Without being limited by a particular theory, a compoundprovided herein (e.g., Compound 4) can affect TAM cell counts in thetumor microenvironment, and can be tested in one or more melanoma modelknown in the art.

Colon Cancer Model

CT26 is an N-nitroso-N-methylurethane-(NNMU) induced, undifferentiatedmouse colon carcinoma cell line. It is cloned to generate the cell linedesignated CT26.WT (ATCC CRL-2638). The syngeneic CT26 colon cancermodel is established by subcutaneous implantation of CT26 cells intoBALB/c mice. This model has been extensively used to study theanti-tumor activity of immunotherapies (Yu et al., Clinical CancerResearch, 2010; Daraiswamy et al., Cancer Research, 2013). This modelcan be used to demonstrate the effects of PI3K inhibitors disclosedherein. In some embodiments, a compound provided herein can beadministered alone or in combination with an anti-PD-L1.

Another colon cancer model is the CR C57BL/6 mice. Female CR C57BL/6mice are set up with 1×106 MC38 tumor cells in 0% Matrigel sc in flank.The cell injection volume is 0.05 ml per mouse. When tumors reached anaverage size of 80-100 mm³, a pair match is made and treatment with acompound provided herein begins.

Glioblastoma Model

Nude mouse xenograft model of human glioma can be used to study theeffects of compounds provided herein in glioblastoma. Tumors aregenerated in athymic nude mice (Taconic Laboratories). The animals areinjected subcutaneously on the right flank with 5×10⁶ U87 human gliomacells in 0.1 ml of PBS supplemented with 0.1% glucose. Tumors aremeasured using an external caliper, every day of treatment, and volumeis calculated. When tumors reaches a volume of 200 mm³, mice arerandomly distributed into different experimental groups and treateddaily with vehicle of the control compound or with blank or a compoundprovided herein at various dosese and schedule. Mice are monitored dailyfor health status and for tumor volumes. After certain days of treatmentmice are sacrified and tumors are removed, measured and weighed. Seee.g., Dolores Hernan Perez de la Ossa, et al., PLOS ONE, 2013, vol. 8(1), e54795.

In some cases, the GL261 glioblastoma model is used. This is a syngeneicglioblastoma multiform model run subcutaneously or orthotopically, andis described in more detail in the examples provided herein. In someembodiments, the model has previously been treated with radiationtherapy but there is a recurrence of the cancer. Compounds providedherein can be administered to models with a recurrence or relapse ofcancer after prior treatment.

Example 228: Microsome Stability Assay

The stability of one or more subject compounds is determined accordingto standard procedures known in the art. For example, stability of oneor more subject compounds is established by an in vitro assay. Forexample, an in vitro microsome stability assay is established thatmeasures stability of one or more subject compounds when reacting withmouse, rat or human microsomes from liver. The microsome reaction withcompounds is performed in 1.5 mL Eppendorf tube. Each tube contains 0.1μL of 10.0 mg/mL NADPH; 75 μL of 20.0 mg/mL mouse, rat or human livermicrosome; 0.4 μL of 0.2 M phosphate buffer, and 425 μL of ddH₂O.Negative control (without NADPH) tube contains 75 μL of 20.0 mg/mLmouse, rat or human liver microsome; 0.4 μL of 0.2 M phosphate buffer,and 525 μL of ddH₂O. The reaction is started by adding 1.0 μL of 10.0 mMtested compound. The reaction tubes are incubated at 37° C. 100 μLsample is collected into new Eppendorf tube containing 300 μL coldmethanol at 0, 5, 10, 15, 30 and 60 minutes of reaction. Samples arecentrifuged at 15,000 rpm to remove protein. Supernatant of centrifugedsample is transferred to new tube. Concentration of stable compoundafter reaction with microsome in the supernatant is measured by LiquidChromatography/Mass Spectrometry (LC-MS).

Example 229: Plasma Stability Assay

The stability of one or more subject compounds in plasma is determinedaccording to standard procedures known in the art. See, e.g., RapidCommun. Mass Spectrom., 10: 1019-1026. The following procedure is anHPLC-MS/MS assay using human plasma; other species including monkey,dog, rat, and mouse are also available. Frozen, heparinized human plasmais thawed in a cold water bath and spun for 10 minutes at 2000 rpm at 4°C. prior to use. A subject compound is added from a 400 μM stocksolution to an aliquot of pre-warmed plasma to give a final assay volumeof 400 μL (or 800 μL for half-life determination), containing 5 μM testcompound and 0.5% DMSO. Reactions are incubated, with shaking, for 0minutes and 60 minutes at 37 C, or for 0, 15, 30, 45 and 60 minutes at37 C for half life determination. Reactions are stopped by transferring50 μL of the incubation mixture to 200 μL of ice-cold acetonitrile andmixed by shaking for 5 minutes. The samples are centrifuged at 6000×gfor 15 minutes at 4° C. and 120 μL of supernatant removed into cleantubes. The samples are then evaporated to dryness and submitted foranalysis by HPLC-MS/MS.

In one embodiment, one or more control or reference compounds (5 μM) aretested simultaneously with the test compounds: one compound,propoxycaine, with low plasma stability and another compound,propantheline, with intermediate plasma stability.

Samples are reconstituted in acetonitrile/methanol/water (1/1/2, v/v/v)and analyzed via (RP)HPLC-MS/MS using selected reaction monitoring(SRM). The HPLC conditions consist of a binary LC pump with autosampler,a mixed-mode, C12, 2×20 mm column, and a gradient program. Peak areascorresponding to the analytes are recorded by HPLC-MS/MS. The ratio ofthe parent compound remaining after 60 minutes relative to the amountremaining at time zero, expressed as percent, is reported as plasmastability. In case of half-life determination, the half-life isestimated from the slope of the initial linear range of the logarithmiccurve of compound remaining (%) vs. time, assuming first order kinetics.

Example 230: Kinase Signaling in Blood

PI3K/Akt/mTOR signaling is measured in blood cells using the phosflowmethod (Methods Enzymol. (2007) 434:131-54). This method is by nature asingle cell assay so that cellular heterogeneity can be detected ratherthan population averages. This allows concurrent distinction ofsignaling states in different populations defined by other markers.Phosflow is also highly quantitative. To test the effects of one or morecompounds provided herein, unfractionated splenocytes, or peripheralblood mononuclear cells are stimulated with anti-CD3 to initiate T-cellreceptor signaling. The cells are then fixed and stained for surfacemarkers and intracellular phosphoproteins. Certain inhibitors providedherein, e.g., PI3K-δ inhibitors, inhibit anti-CD3 mediatedphosphorylation of Akt-S473 and S6, whereas rapamycin inhibits S6phosphorylation and enhances Akt phosphorylation under the conditionstested. Certain inhibitors provided herein, e.g., PI3K-γ inhibitors, acton GPCR ligand (e.g., CCL2, CXCL12, or IL8) stimulation of phospho AKTin blood cells. Accordingly, to test the effect of one or more compoundsprovided herein, unfractionated splenocytes, or peripheral bloodmononuclear cells are contacted with a GPCR ligand. The cells are thenfixed and stained for surface markers and intracellular phosphoproteins.

Similarly, aliquots of whole blood are incubated for 15 minutes withvehicle (e.g., 0.1% DMSO) or kinase inhibitors at variousconcentrations, before addition of stimuli to crosslink the T cellreceptor (TCR) (e.g., anti-CD3 with secondary antibody) or the B cellreceptor (BCR) using anti-kappa light chain antibody (Fab′2 fragments).After approximately 5 and 15 minutes, samples are fixed (e.g., with cold4% paraformaldehyde) and used for phosflow. Surface staining is used todistinguish T and B cells using antibodies directed to cell surfacemarkers that are known to the art. The level of phosphorylation ofkinase substrates such as Akt and S6 are then measured by incubating thefixed cells with labeled antibodies specific to the phosphorylatedisoforms of these proteins. The population of cells are then analyzed byflow cytometry.

Example 231: Colony Formation Assay

Murine bone marrow cells freshly transformed with a p190 BCR-Ablretrovirus (herein referred to as p190 transduced cells) are plated inthe presence of various drug combinations in M3630 methylcellulose mediafor about 7 days with recombinant human IL-7 in about 30% serum, and thenumber of colonies formed is counted by visual examination under amicroscope.

Alternatively, human peripheral blood mononuclear cells are obtainedfrom Philadelphia chromosome positive (Ph+) and negative (Ph−) patientsupon initial diagnosis or relapse. Live cells are isolated and enrichedfor CD19+CD34+ B cell progenitors. After overnight liquid culture, cellsare plated in methocult GF+ H4435 (Stem Cell Technologies), supplementedwith cytokines (IL-3, IL-6, IL-7, G-CSF, GM-CSF, CF, Flt3 ligand, anderythropoietin) and various concentrations of known chemotherapeuticagents in combination with compounds of the present disclosure. Coloniesare counted by microscopy 12-14 days later. This method can be used totest for evidence of additive or synergistic activity.

Example 232: In Vivo Effect of Kinase Inhibitors on Leukemic Cells

Female recipient mice are lethally irradiated from a γ source in twodoses about 4 hr apart, with approximately 5Gy each. About 1 hr afterthe second radiation dose, mice are injected i.v. with about 1×10⁶leukemic cells (e.g., Ph+ human or murine cells, or p190 transduced bonemarrow cells). These cells are administered together with aradioprotective dose of about 5×10⁶ normal bone marrow cells from 3-5week old donor mice. Recipients are given antibiotics in the water andmonitored daily. Mice who become sick after about 14 days are euthanizedand lymphoid organs are harvested for analysis. Kinase inhibitortreatment begins about 10 days after leukemic cell injection andcontinues daily until the mice become sick or a maximum of approximately35 days post-transplant. Inhibitors are given by oral lavage.

Peripheral blood cells are collected approximately on day 10(pre-treatment) and upon euthanization (post treatment), contacted withlabeled anti-hCD4 antibodies and counted by flow cytometry. This methodcan be used to demonstrate that the synergistic effect of one or morecompounds provided herein in combination with known chemotherapeuticagents can reduce leukemic blood cell counts as compared to treatmentwith known chemotherapeutic agents (e.g., Gleevec) alone under theconditions tested.

Example 233: Treatment of Lupus Disease Model Mice

N014181 Mice lacking the inhibitory receptor FcγRIIb that opposes PI3Ksignaling in B cells develop lupus with high penetrance. FcγRIIbknockout mice (R2KO, Jackson Labs) are considered a valid model of thehuman disease as some lupus patients show decreased expression orfunction of FcγRIIb (S. Bolland and J. V. Ravtech 2000. Immunity12:277-285).

The R2KO mice develop lupus-like disease with anti-nuclear antibodies,glomerulonephritis and proteinurea within about 4-6 months of age. Forthese experiments, the rapamycin analogue RAD001 (available from LCLaboratories) is used as a benchmark compound, and administered orally.This compound has been shown to ameliorate lupus symptoms in theB6.Sle1z.Sle3z model (T. Wu et al. J. Clin Invest. 117:2186-2196).

The NZB/W F1 mice that spontaneously develop a systemic autoimmunedisease is a model of lupus. The murine NZB/W F1 lupus model has manyfeatures of human lupus, and is characterized by elevated levels ofanti-nuclear and anti-dsDNA autoantibodies; a critical role forplasmacytoid dendritic cells and IFN-α; T-cell, B-cell, macrophageinvolvement; pheymolytic anemia; progressive immune complexglomerulonephritis; proteinurea; severity and incidence more pronouncedin females; and decreased survival. Treatment with a compound providedherein can be determined by evaluation of urine protein scores, organweights, plasma anti-dsDNA IgG levels, and histopathology of thekidneys. The mice are treated starting at 20 weeks of age for aprofilactic model and at 23 weeks of age for a therapeutic model. Bloodand urine samples are obtained throughout the testing period, and testedfor antinuclear antibodies (in dilutions of serum) or proteinconcentration (in urine). Serum is also tested for anti-ssDNA andanti-dsDNA antibodies by ELISA. Glomerulonephritis is assessed in kidneysections stained with H&E at the end of the study, or survival can be anendpoint. For example, the proteozome inhibitor Bortezimib is effectiveat blocking disease in the NZB/W model in both the profilactic andtherapeutic model with reductions in auto-antibody production, kidneydamage, and improvements in survival (Nature Medicine 14, 748-755(2008)).

Lupus disease model mice such as R2KO, BXSB or MLR/lpr are treated atabout 2 months old, approximately for about two months. Mice are givendoses of: vehicle, RAD001 at about 10 mg/kg, or compounds providedherein at approximately 1 mg/kg to about 500 mg/kg. Blood and urinesamples are obtained throughout the testing period, and tested forantinuclear antibodies (in dilutions of serum) or protein concentration(in urine). Serum is also tested for anti-ssDNA and anti-dsDNAantibodies by ELISA. Animals are euthanized at day 60 and tissuesharvested for measuring spleen weight and kidney disease.Glomerulonephritis is assessed in kidney sections stained with H&E.Other animals are studied for about two months after cessation oftreatment, using the same endpoints.

This established art model can be employed to demonstrate that thekinase inhibitors provided herein can suppress or delay the onset oflupus symptoms in lupus disease model mice.

Example 234: Murine Bone Marrow Transplant Assay

Female recipient mice are lethally irradiated from a γ ray source. About1 hr after the radiation dose, mice are injected with about 1×106leukemic cells from early passage p190 transduced cultures (e.g., asdescribed in Cancer Genet Cytogenet. 2005 August; 161(1):51-6). Thesecells are administered together with a radioprotective dose ofapproximately 5×10⁶ normal bone marrow cells from 3-5 wk old donor mice.Recipients are given antibiotics in the water and monitored daily. Micewho become sick after about 14 days are euthanized and lymphoid organsharvested for flow cytometry and/or magnetic enrichment. Treatmentbegins on approximately day 10 and continues daily until mice becomesick, or after a maximum of about 35 days post-transplant. Drugs aregiven by oral gavage (p.o.). In a pilot experiment, a dose ofchemotherapeutic that is not curative but delays leukemia onset by aboutone week or less is identified; controls are vehicle-treated or treatedwith chemotherapeutic agent, previously shown to delay but not cureleukemogenesis in this model (e.g., imatinib at about 70 mg/kg twicedaily). For the first phase, p190 cells that express eGFP are used, andpostmortem analysis is limited to enumeration of the percentage ofleukemic cells in bone marrow, spleen and lymph node (LN) by flowcytometry. In the second phase, p190 cells that express a tailless formof human CD4 are used and the postmortem analysis includes magneticsorting of hCD4+ cells from spleen followed by immunoblot analysis ofkey signaling endpoints: p Akt-T308 and S473; pS6 and p4EBP-1. Ascontrols for immunoblot detection, sorted cells are incubated in thepresence or absence of kinase inhibitors of the present disclosureinhibitors before lysis. Optionally, “phosflow” is used to detect pAkt-S473 and pS6-S235/236 in hCD4-gated cells without prior sorting.These signaling studies are particularly useful if, for example,drug-treated mice have not developed clinical leukemia at the 35 daytime point. Kaplan-Meier plots of survival are generated and statisticalanalysis done according to methods known in the art. Results from p190cells are analyzed separated as well as cumulatively.

Samples of peripheral blood (100-200 μL) are obtained weekly from allmice, starting on day 10 immediately prior to commencing treatment.Plasma is used for measuring drug concentrations, and cells are analyzedfor leukemia markers (eGFP or hCD4) and signaling biomarkers asdescribed herein.

This general assay known in the art can be used to demonstrate thateffective therapeutic doses of the compounds provided herein can be usedfor inhibiting the proliferation of leukemic cells.

Example 235: Matrigel Plug Angiogenesis Assay

Matrigel containing test compounds are injected subcutaneously orintraocularly, where it solidifies to form a plug. The plug is recoveredafter 7-21 days in the animal and examined histologically to determinethe extent to which blood vessels have entered it. Angiogenesis ismeasured by quantification of the vessels in histologic sections.Alternatively, fluorescence measurement of plasma volume is performedusing fluorescein isothiocyanate (FITC)-labeled dextran 150. The resultsare expected to indicate one or more compounds provided herein thatinhibit angiogenesis and are thus expected to be useful in treatingocular disorders related to aberrant angiogenesis and/or vascularpermeability.

Example 236: Corneal Angiogenesis Assay

A pocket is made in the cornea, and a plug containing an angiogenesisinducing formulation (e.g., VEGF, FGF, or tumor cells), when introducedinto this pocket, elicits the ingrowth of new vessels from theperipheral limbal vasculature. Slow-release materials such as ELVAX(ethylene vinyl copolymer) or Hydron are used to introduce angiogenesisinducing substances into the corneal pocket. Alternatively, a spongematerial is used.

The effect of putative inhibitors on the locally induced (e.g., spongeimplant) angiogenic reaction in the cornea (e.g., by FGF, VEGF, or tumorcells). The test compound is administered orally, systemically, ordirectly to the eye. Systemic administration is by bolus injection or,more effectively, by use of a sustained-release method such asimplantation of osmotic pumps loaded with the test inhibitor.Administration to the eye is by any of the methods described hereinincluding, but not limited to eye drops, topical administration of acream, emulsion, or gel, intravitreal injection.

The vascular response is monitored by direct observation throughout thecourse of the experiment using a stereomicroscope in mice. Definitivevisualization of the corneal vasculature is achieved by administrationof fluorochrome-labeled high-molecular weight dextran. Quantification isperformed by measuring the area of vessel penetration, the progress ofvessels toward the angiogenic stimulus over time, or in the case offluorescence, histogram analysis or pixel counts above a specific(background) threshold.

The results can indicate one or more compounds provided herein inhibitangiogenesis and thus can be useful in treating ocular disorders relatedto aberrant angiogenesis and/or vascular permeability.

Example 237: Microtiter-Plate Angiogenesis Assay

The assay plate is prepared by placing a collagen plug in the bottom ofeach well with 5-10 cell spheroids per collagen plug each spheroidcontaining 400-500 cells. Each collagen plug is covered with 1100 μL ofstorage medium per well and stored for future use (1-3 days at 37° C.,5% CO₂). The plate is sealed with sealing. Test compounds are dissolvedin 200 μL assay medium with at least one well including a VEGF positivecontrol and at least one well without VEGF or test compound as anegative control. The assay plate is removed from the incubator andstorage medium is carefully pipeted away. Assay medium containing thetest compounds are pipeted onto the collagen plug. The plug is placed ina humidified incubator for (37° C., 5% CO₂) 24-48 hours. Angiogenesis isquantified by counting the number of sprouts, measuring average sproutlength, or determining cumulative sprout length. The assay can bepreserved for later analysis by removing the assay medium, adding 1 mLof 10% paraformaldehyde in Hanks BSS per well, and storing at 4° C. Theresults are expected to identify compounds that inhibit angiogenesis invarious cell types tested, including cells of ocular origin.

Example 238: Combination Use of PI3K-δ Inhibitors and Agents thatInhibit IgE Production or Activity

The compounds as provided herein can present synergistic or additiveefficacy when administered in combination with agents that inhibit IgEproduction or activity. Agents that inhibit IgE production include, forexample, one or more of TEI-9874,2-(4-(6-cyclohexyloxy-2-naphtyloxy)phenylacetamide)benzoic acid,rapamycin, rapamycin analogs (i.e., rapalogs), TORC1 inhibitors, TORC2inhibitors, and any other compounds that inhibit mTORC1 and mTORC2.Agents that inhibit IgE activity include, for example, anti-IgEantibodies such as Omalizumab and TNX-901.

One or more of the subject compounds capable of inhibiting PI3K-δ can beefficacious in treatment of autoimmune and inflammatory disorders(AIID), for example, rheumatoid arthritis. If any of the compoundscauses an undesired level of IgE production, one can choose toadminister it in combination with an agent that inhibits IgE productionor IgE activity. Additionally, the administration of PI3K-δ or PI3K-δ/γinhibitors as provided herein in combination with inhibitors of mTOR canalso exhibit synergy through enhanced inhibition of the PI3K pathway.Various in vivo and in vitro models can be used to establish the effectof such combination treatment on AIID including, but not limited to: (a)in vitro B-cell antibody production assay, (b) in vivo TNP assay, and(c) rodent collagen induced arthritis model.

(a) B-Cell Assay

Mice are euthanized, and the spleens are removed and dispersed through anylon mesh to generate a single-cell suspension. The splenocytes arewashed (following removal of erythrocytes by osmotic shock) andincubated with anti-CD43 and anti-Mac-1 antibody-conjugated microbeads(Miltenyi Biotec). The bead-bound cells are separated from unbound cellsusing a magnetic cell sorter. The magnetized column retains the unwantedcells and the resting B cells are collected in the flow-through.Purified B-cells are stimulated with lipopolysaccharide or an anti-CD40antibody and interleukin 4. Stimulated B-cells are treated with vehiclealone or with PI3K-δ inhibitors as provided herein with and without mTORinhibitors such as rapamycin, rapalogs, or mTORC1/C2 inhibitors. Theresults are expected to show that in the presence of mTOR inhibitors(e.g., rapamycin) alone, there is little to no substantial effect on IgGand IgE response. However, in the presence of PI3K-δ and mTORinhibitors, the B-cells are expected to exhibit a decreased IgG responseas compared to the B-cells treated with vehicle alone, and the B-cellsare expected to exhibit a decreased IgE response as compared to theresponse from B-cells treated with PI3K-δ inhibitors alone.

(b) TNP Assay

Mice are immunized with TNP-Ficoll or TNP-KHL and treated with: vehicle,a PI3K-δ inhibitor, an mTOR inhibitor, for example rapamycin, or aPI3K-δ inhibitor in combination with an mTOR inhibitor such asrapamycin. Antigen-specific serum IgE is measured by ELISA using TNP-BSAcoated plates and isotype specific labeled antibodies. It is expectedthat mice treated with an mTOR inhibitor alone exhibit little or nosubstantial effect on antigen specific IgG3 response and nostatistically significant elevation in IgE response as compared to thevehicle control. It is also expected that mice treated with both PI3K-δinhibitor and mTOR inhibitor exhibit a reduction in antigen specificIgG3 response as compared to the mice treated with vehicle alone.Additionally, the mice treated with both PI3K-δ inhibitor and mTORinhibitor exhibit a decrease in IgE response as compared to the micetreated with PI3K-δ inhibitor alone.

(c) Rat Collagen Induced Arthritis Model

Female Lewis rats are anesthetized and given collagen injectionsprepared and administered as described previously on day 0. On day 6,animals are anesthetized and given a second collagen injection. Calipermeasurements of normal (pre-disease) right and left ankle joints areperformed on day 9. On days 10-11, arthritis typically occurs and ratsare randomized into treatment groups. Randomization is performed afterankle joint swelling is obviously established and there is good evidenceof bilateral disease.

After an animal is selected for enrollment in the study, treatment isinitiated. Animals are given vehicle, PI3K-δ inhibitor, or PI3K-δinhibitor in combination with rapamycin. Dosing is administered on days1-6. Rats are weighed on days 1-7 following establishment of arthritisand caliper measurements of ankles taken every day. Final body weightsare taken on day 7 and animals are euthanized.

The combination treatment using a compound as provided herein andrapamycin can provide greater efficacy than treatment with PI3K-δinhibitor alone.

Example 239: Delayed Type Hypersensitivity Model

DTH is induced by sensitizing 60 BALB/c male mice on day 0 and day 1with a solution of 0.05% 2,4 dinitrofluorobenzene (DNFB) in a 4:1acetone/olive oil mixture. Mice are gently restrained while 20 μL ofsolution is applied to the hind foot pads of each mouse. The hind footpads of the mice are used as they represent an anatomical site that canbe easily isolated and immobilized without anesthesia. On day 5, miceare administered a single dose of vehicle, a compound provided herein at10, 3, 1, or 0.3 mg/kg, or dexamethasone at a dose of 5 mg/kg by oralgavage. Thirty minutes later mice are anaesthetized, and a solution of0.25% DNFB in a 4:1 acetone/olive oil solution is applied to the leftinner and outer ear surface. This application results in the inductionof swelling to the left ear and under these conditions, all animalsresponded to this treatment with ear swelling. A vehicle controlsolution of 4:1 acetone/olive oil is applied to the right inner andouter ear. Twenty four hours later, mice are anaesthetized, andmeasurements of the left and right ear are taken using a digitalmicrometer. The difference between the two ears is recorded as theamount of swelling induced by the challenge of DNFB. Drug treatmentgroups are compared to vehicle control to generate the percent reductionin ear swelling. Dexamethasone is routinely used as a positive controlas it has broad anti-inflammatory activity.

Example 240: Peptidoglycan-Polysaccharide Rat Arthritic Model

(a) Systemic Arthritis Model

All injections are performed under anesthesia. 60 female Lewis rats(150-170) are anesthetized by inhalation isoflurane using a small animalanesthesia machine. The animals are placed in the induction chamberuntil anesthetized by delivery of 4-5% isoflurane in O₂ and then held inthat state using a nose cone on the procedure table. Maintenance levelof isoflurane is at 1-2%. Animals are injected intraperitoneally (i.p.)with a single injection of purified PG-PS 10S Group A, D58 strain(concentration 25 μg/g of bodyweight) suspended in sterile 0.85% saline.Each animal receives a total volume of 500 microliters administered inthe lower left quadrant of the abdomen using a 1 milliliter syringe witha 23 gauge needle. Placement of the needle is critical to avoidinjecting the PG-PS 10S into either the stomach or caecum. Animals areunder continuous observation until fully recovered from anesthesia andmoving about the cage. An acute response of a sharp increase in anklemeasurement, typically 20% above baseline measurement can peak in 3-5days post injection. Treatment with test compounds can be PO, SC, IV orIP. Rats are dosed no more than two times in a 24 hour time span.Treatment can begin on day 0 or any day after that through day 30. Theanimals are weighed on days 0, 1, 2, 3, 4, 5, 6, 7 and beginning againon day 12-30 or until the study is terminated. Paw/ankle diameter ismeasured with a digital caliper on the left and right side on day 0prior to injection and again on day 1, 2, 3, 4, 5, 6 and 7. On day 12,measurements begin again and continue on through day 30. At this time,animals can be anesthetized with isoflurane, as described above, andterminal blood samples can be obtained by tail vein draws for theevaluation of the compound blood levels, clinical chemistry orhematology parameters. Animals are then euthanized with carbon dioxideoverdose. A thoracotomy can be conducted as a means of deathverification.

(B) Monoarticular Arthritis Model

All injections are performed under anesthesia. 60 female Lewis rats(150-170) are anesthetized by inhalation isoflurane using a small animalanesthesia machine. The animals are placed in the induction chamberuntil anesthetized by delivery of 4-5% isoflurane in O₂ and then held inthat state using a nose cone on the procedure table. Maintenance levelof isoflurane is at 1-2%. Animals are injected intra-articular (i.a.)with a single injection of purified PG-PS 100P Group A, D58 strain(concentration 500 μg/mL) suspended in sterile 0.85% saline. Each ratreceives a total volume of 10 microliters administered into thetibiotalar joint space using a 1 milliliter syringe with a 27 gaugeneedle. Animals are under continuous observation until fully recoveredfrom anesthesia and moving about the cage. Animals that respond 2-3 dayslater with a sharp increase in ankle measurement, typically 20% abovebaseline measurement on the initial i.a. injection, are included in thestudy. On day 14, all responders are anesthetized again using theprocedure previously described. Animals receive an intravenous (I.V.)injection of PG-PS (concentration 250 μL/mL). Each rat receives a totalvolume of 400 microliters administered slowly into the lateral tail veinusing a 1 milliliter syringe with a 27 gauge needle. Baseline anklemeasurements are measured prior to IV injection and continue through thecourse of inflammation or out to day 10. Treatment with test compoundswill be PO, SC, IV or IP. Rats are dosed no more than two times in a 24hour time span. Treatment can begin on day 0 or any day after thatthrough day 24. The animals are weighed on days 0, 1, 2, 3, 4, 5, andbeginning again on day 14-24 or until the study is terminated. Paw/anklediameter is measured with a digital caliper on the left and right sideon day 0 prior to injection and again on day 1, 2, 3, 4, 5, andbeginning again on day 14-24 or until the study is terminated. At thistime, animals can be anesthetized with isoflurane, as described above,and terminal blood samples can be obtained by tail vein draws for theevaluation of the compound blood levels, clinical chemistry orhematology parameters. Animals are them euthanized with carbon dioxideoverdose. A thoracotomy can be conducted as a means of deathverification.

Example 241: Mice Models for Asthma

Efficacy of a compound provided herein in treating, preventing and/ormanaging asthma can be assessed using an conventional animal modelsincluding various mice models described in, for example, Nials et al.,Dis Model Mech. 1(4-5): 213-220 (2008).

(a) Acute Allergen Challenge Models

Several models are known in the art and any of such models can be used.Although various allergens can be used to induce asthma-like conditions,the principle is consistent throughout the methods. Briefly, asthma-likeconditions are induced through multiple systemic administration of theallergen (e.g., ova, house dust mite extracts and cockroach extracts) inthe presence of an adjuvant such as aluminum hydroxide. Alternatively,an adjuvant-free system can be used, but it usually requires a highernumber of exposures to achieve suitable sensitization. Once induced,animals exhibit many key features of clinical asthma such as: elevatedlevels of IgE; airway inflammation; goblet cell hyperplasia; epithelialhypertrophy; AHR ro specific stimuli; and early and late phasebronchoconstriction. Potential efficacy of a compound thus can beassessed by determining whether one or more of these clinical featuresare reversed or mitigated.

(b) Chronic Allergen Challenge Models

Chronic allergen challenge models aim to reproduce more of the featuresof the clinical asthma, such as airway remodeling and persistent AHR,than acute challenge models. While allergens similar to those used inacute allergen challenge models can be used, in chronic allergenchallenge models, animals are subjected to repeated exposure of theairways to low levels of allergen for a period of up to 12 weeks. Onceinduced, animals exhibit key features of human asthma such as:allergen-dependent sensitization; a Th2-dependent allergic inflammationcharacterized by eosinophillic influx into the airway mucosa; AHR; andairway remodeling as evidenced by goblet cell hyperplasia, epithelialhypertrophy, subepithelial or peribronchiolar fibrosis. Potentialefficacy of a compound thus can be assessed by determining whether oneor more of these clinical features are reversed or mitigated.

Example 242: Models for Psoriasis

Efficacy of a compound provided herein in treating, preventing and/ormanaging psoriasis can be assessed using an conventional animal modelsincluding various animal models described in, for example, Boehncke etal., Clinics in Dermatology, 25: 596-605 (2007).

As an example, the mouse model based on adoptive transfer ofCD4⁺CD45RB^(hi) T cells described in Hong et al., J. Immunol., 162:7480-7491 (1999) can be made. Briefly, female BALB/cBY (donor) andC.B.-17/Prkdc scid/scid (recipient) mice are housed in a specificpathogen-free environment and are used between 6 and 8 weeks of age.CD4⁺ T cells are enriched from BALB/cBy splenocytes using a mouse CD4enrichment kit. The cells are then labeled with PE-conjugated anti-CD4,FITC-conjugated anti-CD45RB, and APC-conjugated anti-CD25 antibodies.Cells are sorted using a cell sorter. CD4⁺CD45RB^(hi)CD25 cells arecollected. Cells are resuspended in saline and 4×10⁸ cells/mouse areinjected i.p. into C.B.-17/Prkdc scid/scid mice. Mice may be dosed withLPS, cytokines, or antibodies as necessary. Mice are monitored forexternal signs of skin lesions twice each week. After the termination,ear, back skin, lymph nodes and spleen may be collected for further exvivo studies.

Example 243: Models for Scleroderma

A compound's efficacy in treating scleroderma can be tested using animalmodels. An exemplary animal model is a mouse model for sclerodermainduced by repeated local injections of bleomycin (“BLM”) described, forexample, in Yamamoto et al., J Invest Dermatol 112: 456-462 (1999), theentirety of which is incorporated herein by reference. This mouse modelprovides dermal sclerosis that closely resembles systemic sclerosis bothhistologically and biochemically. The sclerotic changes observed in themodel include, but are not limited to: thickened and homogenous collagenbundles and cellular filtrates; gradual increase in number of mastcells; degranulation of mast cells; elevated histamine release; increasein hydroxyproline in skin; presence of anti-nuclear antibody in serum;and strong expression of transforming growth factor β-2 mRNA. Therefore,efficacy of a compound in treating scleroderma can be assessed bymonitoring the lessening of one or more of these changes.

Briefly, the following exemplary procedures can be used to generate themouse model for scleroderma: Specific pathogen-free, female BALB/C miceand C3H mice of 6 weeks old, weighing about 20 g, are purchased andmaintained with food and water ad libitum. BLM is dissolved in PBS atdiffering concentrations and sterilized with filtration. Aliquots ofeach concentration of BLM or PBS are injected subcutaneously into theshaved back of the mice daily for 1-4 weeks with a needle.Alternatively, mice are injected every other day.

Histolopathological and biochemical changes induced can be assessedusing any methods commonly practiced in the field. For example,histopathological changes can be assessed using a standardavidine-biotin peroxidase technique with anti-L3T4 monoclonal antibody,anti-Lyt2 monoclonal antibody, anti-mouse pan-tissue-fixed macrophageantibody, anti-stem cell factor monoclonal antibody, anti-transforminggrowth factor-β polyclonal antibody, and anti-decorin antibody. Cytokineexpression of cellular infiltrates can be assessed by using severalanti-cytokine antibodies. Hydroxyproline level can be assessed byhydrolyzing skin pieces with hydrochloric acid, neutralizing with sodiumhydroxide, and colorimetrically assessing the hydrolates at 560 nm withp-dimethylaminobenzaldehyde. Pepsin-resistant collagen can be assessedby treating collagen sample extracted from biopsied tissues andanalyzing by polyacrylamide stacking gel electrophoresis. Mast cells canbe identified by toluidine blue, and cells containing matachromaticgranules can be counted under high magnification of a light microscope.Serum levels of various cytokines can be assessed by enzyme-linkedimmunosorbent assay, and mRNA levels of the cytokines can be assessed byreverse-transcriptase polymerase chain reaction. Autoantibodies in serumcan be detected using 3T3 fibroblasts as the substrate for thescreening.

Example 244: Models for Myositis

A compound's efficacy in treating myositis (e.g., dermatomyositis) canbe tested using animal models known in the art. One such example is thefamilial canine dermatomyositis model described in Hargis et al., AJP120(2): 323-325 (1985). Another example is the rabbit myosin inducedmouse model described in Phyanagi et al., Arthritis & Rheumatism,60(10): 3118-3127 (2009).

Briefly, 5-week old male SJL/J mice are used. Purified myosin fromrabbit skeletal muscle (6.6 mg/ml) is emulsified with an equal amount ofFreund's complete adjuvant and 3.3 mg/ml Mycobacterium butyricum. Themice are immunized repeatedly with emulsified rabbit myosin. Oncemyositis is induced, inflammatory cell filtration and necrotic musclefiber should be evident in the model. In the muscles of animals, CD4⁺ Tcells are mainly located in the perimysum and CD8⁺ T cells are mainlylocated in the endomysium and surround non-necrotic muscle fibers. TNFα,IFNγ and perforin are upregulated and intercellular adhesion molecule 1is increased in the muscles.

To assess the efficacy of a compound, following administration of thecompound through adequate route at specified dose, the mice are killedand muscle tissues are harvested. The muscle tissue is immediatelyfrozen in chilled isopentane precooled in liquid nitrogen, and thencryostat sections are prepared. The sections are stained withhematoxylin and eosin for counting of number of infiltrated cells. Threesections from each mouse are prepared and photomicrographs are obtained.For immunohistochemical tests, cryostat sections of muscle are dried andfixed in cold acetone at −20° C. The slides are rehydrated in PBS, andthen endogeneous peroxide activity is blocked by incubation in 1%hydrogen peroxide. The sections are incubated overnight with ratanti-mouse CD4 monoclonal antibody, rat anti-mouse CD8 monoclonalantibody, rat anti-mouse F4/80 monoclonal antibody or normal rat IgG inantibody diluent. The samples are washed with PBS and incubated withbiotin-conjugated rabbit anti-rat IgG pretreated with 5% normal mouseserum. After washing with PBS, the samples are incubated withstreptavidin-horseradish peroxidase. After washing PBS, diaminobenzidineis used for visualization.

Example 245: Models for Sjögren Syndrome

A compound's efficacy in treating Sjögren's syndrome can be tested usinganimal models known in the art, for example, those described in Chioriniet al., Journal of Autoimmunity 33: 190-196 (2009). Examples include:mouse model spontaneously developed in first filial generation of NZBmice crossed to NZW mice (see, e.g., Jonsson et al., Clin ImmunolImmunopathol 42: 93-101 (1987); mouse model induced by i.p. injection ofincomplete Freund's adjuvant (id.; Deshmukh et al., J Oral Pathol Med38: 42-27 (2009)); NOD mouse models wherein Sjögren's phenotype isdeveloped by specific genotypes (see, e.g., Cha et al., Arthritis Rheum46: 1390-1398 (2002); Kong et al., Clin Exp Rheumatol 16: 675-681(1998); Podolin et al., J Exp Med 178: 793-803 (1993); and Rasooly etal., Clin Immunol Immunopathol 81: 287-292 (1996)); mouse modeldeveloped in spontaneous lpr mutation; mouse model developed in Id3knock-out mice (see, e.g., Li et al., Immunity 21: 551-560 (2004));mouse model developed in PI3K knock-out mice (see, e.g., Oak et al.,Proc Natl Acad Sci USA 103: 16882-16887 (2006)); mouse model developedin BAFF over-expressing transgenic mice (see, e.g., Groom et al., J ClinInvest 109: 59-68 (2002)); mouse model induced by injection of Roantigen into BALB/c mice (see, e.g., Oh-Hora et al., Nat. Immunol 9:432-443 (2008)); mouse model induced by injection of carbonic anhydraseII (see, e.g., Nishimori et al., J Immunol 154: 4865-4873 (1995); mousemodel developed in IL-14 over-expressing transgenic mice (see, e.g.,Shen et al., J Immunol 177: 5676-5686 (2006)); and mouse model developedin IL-12 expressing transgenic mice (see, e.g., McGrath-Morrow et al.,Am J Physiol Lung Cell Mol Physiol 291: L837-846 (2006)).

Example 246: Models for Immune Complex Mediated Disease

The Arthus reaction is a type 3 immune response to immune complexes, andthus, can be a mechanistic model supporting therapeutic hypothesis forimmune complex mediated diseases such as rheumatoid arthritis, lupus andother autoimmune diseases. For example, PI3Kγ and δ deficient mice canbe used as experimental models of the Arthus reaction and provideassessment of therapeutic potential of a compound as to the treatment ofimmune complex mediated diseases. The Arthus reaction can be inducedusing the following exemplary procedures as described in Konrad et al.,Journal of Biological Chemistry (2008 283(48): 33296-33303.

PI3Kγ- and PI3Kδ-deficient mice are maintained under dry barrierconditions. Mice are anesthetized with ketamine and xylazine, and thetrachea is cannulated. Appropriate amount of protein G-purified anti-OVAIgG Ab is applied, and appropriate amount of OVA antigen is givenintravenously. For PI3K blocking experiments, wortmanin is givenintratracheally together with the application of anti-OVA igG. Mice arekilled at 2-4 hours after initiation of inflammation, and desired followup assessments can be performed using methods known in the art.

Example 247: PI3-Kinase Promega™ Assay

Promega ADP-Glo Max assay kit (Cat. No. V7002) was utilized to determineIC₅₀ values for α, β, δ and γ isoforms of human Class I PI3 kinases(Millipore). Samples of kinase (20 nM α or δ, 40 nM β or γ isoform) wereincubated with compound for 15 minutes at room temperature in reactionbuffer (15 mM HEPES pH 7.4, 20 mM NaCl, 1 mM EGTA, 0.02% Tween 20, 10 mMMgCl₂, 0.2 mg/mL bovine-γ-globulins) followed by addition ofATP/diC8-PtdInsP mixture to give final concentrations of 3 mM ATP and500 uM diC₈-PtdInsP. Reactions were incubated at room temperature for 2hours followed by addition of 25 uL of stop solution. After a 40-minuteincubation at room temperature, 50 uL of Promega detection mix was addedfollowed by incubation for 1 hour at room temperature. Plates were thenread on Envision plate reader in luminescence mode. Data was convertedto % inhibition using the following equation below:

${\%\mspace{14mu}{inhibition}} = {100 - ( {\lbrack \frac{S - {Pos}}{{Neg} - {Pos}} \rbrack*100} )}$where S is the sample luminescence, Pos is a positive control withoutadded PI3K, Neg is the negative control without added compound. Data wasthen plotted as % inhibition vs compound concentration. Data fit to 4parameter logistic equation to determine IC₅₀ values:

${\%\mspace{14mu}{Inhibition}} = \frac{\max - \min}{1 - ( \frac{{IC}_{50}^{h}}{\lbrack I\rbrack^{h}} )}$

Certain compounds provided herein were tested in PI3-Kinase PromegaAssay using procedures as described above to determine IC₅₀ values forα, β, δ and/or γ isoforms. The IC₅₀ values are summarized in Table 15.

Example 248: Isoform-Selective Cellular Assays

(a) PI3K-δ Selective Assay

A compound's ability in selectively inhibiting PI3K-δ can be assessedusing RAJI cells, i.e., B lymphocyte cells derived from lymphomapatients. Briefly, serum-starved RAJI cells are stimulated withanti-human IgM, thereby causing signaling through the B-cell receptors,as described in, for example, He et al., Leukemia Research (2009) 33:798-802. B-cell receptor signaling is important for the activation,differentiation, and survival of B cells and certain B-cell derivedcancers. Reduction of phospho-AKT is indicative of compounds that mayinhibit B-cell proliferation and function in certain diseases. Bymonitoring the reduction of phospho-AKT in stimulated RAJI cells (usingfor example, phospho-AKT antibodies), a compound's potential efficacy inselectively inhibiting PI3Kδ can be assessed.

Certain compounds provided herein were tested in RAJI cell model usingprocedures as described above. The IC₅₀ values for phospho-AKT aresummarized in Table 15.

(b) PI3K-γ Selective Assay

A compound's ability in selectively inhibiting PI3K-γ can be assessedusing RAW264.7 macrophages. Briefly, serum-starved RAW264.7 cells arestimulated with a known GPCR agonist C5a. See, e.g., Camps et al.,Nature Medicine (2005) 11(9):936-943. Cells can be treated with testcompounds prior to, simultaneously with, or subsequent to thestimulation by C5a. RAW 264.7 cells respond to the complement componentfragment C5a through activation of the C5a receptor, and the C5areceptor activates macrophages and induces cell migration. Testcompounds' ability to inhibit C5a-mediated AKT phosphorylation isindicative of selective inhibition of PI3K-γ. Thus, by monitoring thereduction of phospho-AKT in stimulated RAW 264.7 cells (using forexample, phospho-AKT antibodies), a compound's potential efficacy inselectively inhibiting PI3Kγ can be assessed.

Certain compounds provided herein were tested in RAW 264.7 cell modelusing procedures as described above. The IC₅₀ values for phospho-AKT aresummarized in Table 15.

(c) PI3K-α Selective Assay

A compound's ability in selectively inhibiting PI3K-α can be assessedusing SKOV-3 cells, i.e., human ovarian carcinoma cell line. Briefly,SKOV-3 cells, in which mutant PI3Kα is constitutively active, can betreated with test compounds. Test compounds' ability to inhibit AKTphosphorylation in SKOV-3 cells, therefore, is indicative of selectiveinhibition of PI3Kα. Thus, by monitoring the reduction of phospho-AKT inSKOV-3 cells (using for example, phospho-AKT antibodies), a compound'spotential efficacy in selectively inhibiting PI3Kα can be assessed.

(d) PI3K-β Selective Assay

A compound's ability in selectively inhibiting PI3K-β can be assessedusing 786-0 cells, i.e., human kidney carcinoma cell line. Briefly,786-O cells, in which PI3Kβ is constitutively active, can be treatedwith test compounds. Test compounds' ability to inhibit AKTphosphorylation in 786-O cells, therefore, is indicative of selectiveinhibition of PI3Kβ. Thus, by monitoring the reduction of phospho-AKT in786-O cells (using for example, phospho-AKT antibodies), a compound'spotential efficacy in selectively inhibiting PI3Kβ can be assessed.

Example 249: Models for Chronic Lymphocytic Leukemia

The TCL-1 transgenic mouse model is a model of chronic lymphocyticleukemia (CLL) that is generated by introducing the human TCL1 geneunder control of a B cell-specific Ig promoter. TCL1 transgenic micedevelop a monoclonal B-cell lymphocytosis that is very similar to humanCLL in immunophenotypic and clinical features. Leukemic splenocytes fromdiseased mice can also be adoptively transferred into syngeneic C57BL/6mice, creating a model allowing for a large number of animals withsynchronously established disease. See e.g., Johnson, A. J., et al.,Blood 108(4):1334-8 (2006).

Placement of 0-1 under the control of a B-cell-specific IgVH promoterand IgH-E enhancer results in a similar B-cell phenotype in which micedevelop normally into adulthood, but then develop enlarged spleens,livers, and lymph nodes associated with high blood lymphocyte counts.The transformed lymphocytes from the TCL-1 mice are GO-1 arrested,clonal, and express CD19+/CD5+/IgM+, as seen in human CLL. Onetransgenic leukemic spleen is harvested from a previously implantedC57BL/6 mouse. Under aseptic conditions, the spleen is dissociated andred blood cells are removed. 2×10⁶ TCL1 leukemic spleen lymphocytes aretransplanted intravenously into 6-8 week old female C57BL/6 recipientmice. Starting two weeks post engraftment, mice are bled bysubmandibular bleed twice weekly to check peripheral blood for percentleukemia. Blood samples are evaluated by flow cytometry for coexpressionof CD5 and CD19 and disease is reported as a percentage of doublepositive cells of the parent CD45+ population. Once the animal hasreached 10-20% double positive staining in peripheral blood, it isassigned to a treatment group. Animals are weighed twice a week andpalpated for splenomegaly. At the end of the study, animals areeuthanized and tissues are harvested for measuring spleen weight andsystemic organ disease. Organ disease is assessed in tissue sectionsstained with H&E. In exemplary studies, a compound provided herein isadministered alone or in combination with a PI3Kδ inhibitor. In oneexemplary study, a compound provided herein is a PI3Kγ inhibitor. Inanother exemplary study, the PI3Kδ inhibitor has a gamma/deltaselectivity ratio of greater than about 50. In yet another exemplarystudy, the PI3Kγ inhibitor has a delta/gamma selectivity ratio ofgreater than about 50.

Example 250: Effect on the Collagen Induced Arthritis Model

Rat collagen arthritis is an experimental model of polyarthritis thathas been widely used for preclinical testing of numerous anti-arthriticagents that are either under preclinical or clinical investigation orare currently used as therapeutics in this disease. The hallmarks ofthis model are reliable onset and progression of robust, easilymeasurable, polyarticular inflammation, marked cartilage destruction inassociation with pannus formation and mild to moderate bone resorptionand periosteal bone proliferation.

Compound BB (PI3K gamma selective compound disclosed here) wasadministered to rats model of collagen induced arthritis (CIA). CompoundBB is an exemplary PI3K gamma selective compound provided herein thathas a delta/gamma selectivity ratio of greater than about 50. CompoundBB was administered at 0.5 mg/kg, 1.5 mg/kg, and 5 mg/kg in groups asidentified below:

TABLE 17 Group Treatment N 1 veh 15 2 Compound BB at 5 mg/kg 11 3Compound BB at 1.5 mg/kg 11 4 Compound BB at 1 mg 11 5 naive 5

Scoring of Joints: collagen arthritic ankles are given scores of 0-5according to the following criteria:

For inflammation, 0=Normal; 0.5=Minimal focal inflammation; 1=Minimalinfiltration of inflammatory cells in synovium/periarticular tissue;2=Mild infiltration; 3=Moderate infiltration with moderate edema;4=Marked infiltration with marked edema; and 5=Severe infiltration withsevere edema.

For pannus, 0=Normal; 0.5=Minimal infiltration of pannus in cartilageand subchondral bone, affects only marginal zones and affects only a fewjoints; 1=Minimal infiltration of pannus in cartilage and subchondralbone, primarily affects marginal zones; 2=Mild infiltration (<¼ of tibiaor tarsals at marginal zones); 3=Moderate infiltration (¼ to ⅓ of tibiaor small tarsals affected at marginal zones); 4=Marked infiltration (½-¾of tibia or tarsals affected at marginal zones); and 5=Severeinfiltration (>¾ of tibia or tarsals affected at marginal zones, severedistortion of overall architecture).

For cartilage damage (emphasis on small tarsals), 0=Normal; 0.5=Minimaldecrease in T blue staining, affects only marginal zones and affectsonly a few joints; 1=Minimal=minimal to mild loss of toluidine bluestaining with no obvious chondrocyte loss or collagen disruption;2=Mild=mild loss of toluidine blue staining with focal mild(superficial) chondrocyte loss and/or collagen disruption;3=Moderate=moderate loss of toluidine blue staining with multifocalmoderate (depth to middle zone) chondrocyte loss and/or collagendisruption, smaller tarsals affected to ½-¾ depth with rare areas offull thickness loss; 4=Marked=marked loss of toluidine blue stainingwith multifocal marked (depth to deep zone) chondrocyte loss and/orcollagen disruption, 1 or 2 small tarsals surfaces have full thicknessloss of cartilage; and 5=Severe=severe diffuse loss of toluidine bluestaining with multifocal severe (depth to tide mark) chondrocyte lossand/or collagen disruption affecting more than 2 cartilage surfaces.

For bone resorption, 0=Normal; 0.5=Minimal resorption affects onlymarginal zones and affects only a few joints; 1=Minimal, small areas ofresorption, not readily apparent on low magnification, rare osteoclasts;2=Mild, more numerous areas of resorption, not readily apparent on lowmagnification, osteoclasts more numerous, <¼ of tibia or tarsals atmarginal zones resorbed; 3=Moderate, obvious resorption of medullarytrabecular and cortical bone without full thickness defects in cortex,loss of some medullary trabeculae, lesion apparent on low magnification,osteoclasts more numerous, ¼ to ⅓ of tibia or tarsals affected atmarginal zones; 4=Marked, full thickness defects in cortical bone, oftenwith distortion of profile of remaining cortical surface, marked loss ofmedullary bone, numerous osteoclasts, ½-¾ of tibia or tarsals affectedat marginal zones; 5=Severe, full thickness defects in cortical bone,often with distortion of profile of remaining cortical surface, markedloss of medullary bone, numerous osteoclasts, >¾ of tibia or tarsalsaffected at marginal zones, severe distortion of overall architecture.

Periosteal New Bone Formation (measure on 16×): Studies that go beyondthe acute inflammatory stage often show varying degrees of periostealnew bone formation. In order to convey the degree of periosteal new boneformation the following scores are applied based on the distribution ofperiosteal bone proliferation and a measurement of width of periostealnew bone formation at the widest location. 0=Normal, no periostealproliferation; 0.5=Minimal focal or multifocal proliferation, measuresless than 127 μm width (1-2) at any location; 1.0=Minimal multifocalproliferation, width at any location measures 127 μm-252 μm (3-4 units);2.0=Mild multifocal on tarsals, diffuse in some locations, width at anylocation 253 μm-441 μm (5-7 units); 3.0=Moderate multifocal on tarsals,diffuse in most other locations, width at any location measures 442μm-630 μm (8-10 units); 4.0=Marked multifocal on tarsals, diffuse atmost other locations, width at any location measures 630 μm-819 μm(11-13 units); 5.0=Severe, multifocal on tarsals, diffuse at most otherlocations, width at any location measures>819 μm (>13 units).

Data was analyzed using a one-way ANOVA or Kruskal-Wallis(non-parametricANOVA) test, along with an appropriate multiplecomparisons post-test. Unless indicated, Bolder BioPATH, Inc. performsstatistical analysis on raw (untransformed) data only. Statistical testsmake certain assumptions regarding the data's normality and homogeneityof variance, and further analysis may be required if testing resulted inviolations of these assumptions. Significance for all tests was set atp≤0.05.

FIG. 1 shows that Compound BB has therapeutic effects in the CIA ratmodel where the mean ankle diameter in inches is measured from day 9 today 17. FIG. 2 shows the individual histopathology scores forinflammation, pannus, cartilage damage, bone resorption, and periostealbone formation. More particularly, in the experiment shown in FIG. 2,inflammation was reduced by 57%, 36%, and 27% when the animals weretreated with Compound BB at 5, 1.5, and 0.5 mg/kg, respectively. Pannuswas reduced by 71%, 44%, and 28% when the animals were treated withCompound BB at 5, 1.5, and 0.5 mg/kg, respectively. Cartilage damage wasreduced by 59%, 45%, and 28% when the animals were treated with CompoundBB at 5, 1.5, and 0.5 mg/kg, respectively. Bone resorption was reducedby 65%, 44%, and 25% when the animals were treated with Compound BB at5, 1.5, and 0.5 mg/kg, respectively. Periosteal bone formation wasreduced by 82%, 52%, and 52% when the animals were treated with CompoundBB at 5, 1.5, and 0.5 mg/kg, respectively. FIG. 3 shows the periostealbone measure. Animals treated with 5 mg/kg of Compound BB hadsignificant 57-82% reductions in all scores resulting in a significant64% reduction in summed scores. Periosteal bone widths were alsosignificantly reduced by 82%. Animals treated with 1.5 mg/kg of CompoundBB had significantly reduced inflammation and cartilage damage (45%)scores, contributing to a significant 42% reduction in summed scores.Although periosteal bone scores were not significantly reduced,periosteal bone widths were significantly reduced by 50%. Animalstreated with 0.5 mg/kg of Compound BB had significantly reducedperiosteal bone widths (47%). Scored parameters were non-significantlyreduced by 27-52%, with a 31% non-significant reduction in summedscores. Results of this study indicate that treatment with Compound BBhad beneficial effects on lesions of collagen-induced arthritis in ratsthat were generally significant at 1.5 mg/kg and above. The ED₅₀ valuefor this treatment was 2.132 mg/kg. All doses of Compound BB testedsignificantly reduced periosteal bone formation.

Example 251: Effect on the IL-8 Driven Murine Air Pouch Model

The purpose is to study the dose response evaluation of the effect ofinfiltrating neutrophils in mouse air pouches stimulated withrecombinant hIL-8 after treatment with Compound BB or Compound AA.Compound AA is a PI3K delta selective compound that has a gamma/deltaselectivity ratio of greater than about 50. Compound BB is a PI3K gammaselective compound described herein that has a delta/gamma selectivityratio of greater than about 50. Balb/c male mice (Stock 000651) wereordered at 6 weeks of age from Jackson Labs/n=60.

Model/Procedure:

The air pouch model is used as a pseudo-synovial space to investigatethe role of specific cytokines and/or effect of compounds oninflammatory mediated events. The pouches are stimulated withrecombinant human IL-8, which is a proinflammatory CXC chemokine thatcan signal through the CXCR1 and CXCR2 receptors. IL-8 chemoattracts andactivates neutrophils.

Pouches are developed by anesthetizing mice using isoflurane at a 3-4%flow. Air pouches are produced by administration of a subcutaneousinjection of 5 mL of sterile air. This can be done by using a 5 mLsyringe with a 22 microfilter attached and a 25G ⅝″ needle attached tothe filter. 5 mLs of air will be drawn into the syringe, the filter withneedle will be placed onto the syringe. The intrascapular area of theback may be swabbed with a 70% alcohol pad. Grabbing the skin betweenthe intrascapular area, a tent will be created by which to insert theneedle subcutaneously between the shoulders of the mouse. Air isinjected slowly, using ones left hand to guide the air to form a longcylindrical pouch (approximately 10 sec), taking precaution not to havethe air slip down the arms or over the head of the mouse. After all 5 mLhas been administered, the needle is withdrawn and the injection site ispinched for approximately 5 sec. Mice are then returned to their cagesand monitored for up to an hour during recovery. Mice are checkedthroughout the day on the day pouches are formed and checked daily forthe next 3 days. On Day 3, mice will be re-injected with 3 ml of sterileair, following the procedure listed above, using a 3 ml syringe with 25Gneedle.

On Day 6, mice are weighed and assigned to a group and compound will bethen be administered. One hour post dose, mice are bled viasubmandibular bleed. Approximately 150 uL of blood is collected into amicrotainer with EDTA. Blood samples are placed on ice, centrifuged for10 min 10,000 RMP at 4 degree. Plasma is placed in an eppendorf tube andstored in the −80 until analyzed by the PK group. Immediately followingblood collection, mice are anesthetized and 1 mL of cold IL-8 stimulantor PBS is injected into the pouch using a 1 mL syringe with a 25Gneedle. The pouch is then massaged gently to ensure the entire lining ofthe pouch has been exposed to the solution.

Sample Collection:

Four hours post stimulation, mice are euthanized by CO₂. Mice are bledvia cardiac puncture and blood is collected in the same manner statedabove. Pouch exudates are then collected in the following manner-3 mlsof cold endotoxin free PBS w/1 mM EDTA is injected into the pouch usinga 3 ml syringe with 20G needle. Pouches are massaged to adequatelysuspend the cells in the pouch, making sure the needle is kept in thepouch to avoid leakage. 2 mL of lavage fluid is drawn out of the pouchand placed in a 5 mL BD Falcon round bottom tube and set on ice untilanalysis. Lavage fluid will be analyzed on the Cell Dyn and celldifferential results will be plotted, with the focus being on totalneutrophil counts from each lavage sample.

Example time line of experimental procedure is listed in the tablebelow:

TABLE 18 Day 0 Day 1 Day 2 Day 3 Day 4 Day 5 Day 6 Animals Daily DailyAnimals Daily Daily Animals weighed weighed monitoring monitoringweighed monitoring monitoring Injection of test Animals Animalscompounds receive 1st receive 2nd 1 h post dose PK injection ofinjection of blood draws air air Administration of stimulant 4 hourspost stimulation animals euthanized Collect terminal PK sample, wash andcollect lavage fluid

For the pouch injections, no stimulation models are injected 1 mLendotoxin free PBS; and stimulation models are injected bug IL-8 in 1 mLof endotoxin free PBS. The reagents are Recombinant Human CXCL8/IL-8-R&DSystems, Cat #208-IL-050/CF Lot # BA3313051; Endotoxin free PBS-Teknova,Cat # P0300; and EDTA (0.5M)-Sigma, Cat # E-7889EDTA (0.5M)-Sigma, Cat #E-7889. Each vial contains 100 ug of IL-8-6 may be used for thisexperiment. Stock is reconstituted to 0.1 mg/mL by adding 1 mL ofendotoxin free PBS, 10 ug/pouch (5.5 mL stock+50 mL PBS). EDTA isprepared using a stock solution concentration of 0.5M and adding 500 ulof stock solution to 250 mL PBS for final concentration of 1 mM EDTA.

Compound BB and Compound AA were tested in the murine air pouch modelaccording to the procedures described above. Compound BB (PI3Kgamma-selective compound) was administered to IL-8 induced neutrophilmodel at 0.1 mg/kg, 0.3 mg/kg, 1 mg/kg, and 3 mg/kg. Compound AA (PI3Kdelta-selective compound) was administered to the IL-8 inducedneutrophil model at 10 mg/kg, 25 mg/kg, and about 50 mg/kg. The resultswere compared to the vehicle. FIG. 4 shows that Compound BB (PI3Kgamma-selective compound) blocks IL-8 induced neutrophil migration, andFIG. 5 shows that Compound AA (PI3K delta-selective compound) does notinhibit IL-8 induced neutrophil migration. This example demonstratesthat PI3K gamma selective compounds provided herein can be used toreduce and/or prevent inflammation.

Example 252: Gamma Selective Compounds Exhibit Synergy with DeltaSelective Compounds

The purpose of this study is to evaluate the anti-cancer effects ofcombinations of PI3K gamma selective compounds provided herein and PI3Kdelta selective compounds. PI3K gamma selective compounds providedherein are combined with a compound that selectively inhibits the PI3Kdelta isoform over the gamma isoform (e.g. delta selective compounds) toprovide synergy. Specifically, compounds provided herein that have adelta/gamma selectivity ratio of greater than about 1 to <10, greaterthan about 10 to <50, or greater than about 50 to <350 are combined witha compound that has a gamma/delta selectivity ratio of greater than afactor of about 1, greater than a factor of about 2, greater than afactor of about 3, greater than a factor of about 5, greater than afactor of about 10, greater than a factor of about 50, greater than afactor of about 100, greater than a factor of about 200, greater than afactor of about 400, greater than a factor of about 600, greater than afactor of about 800, greater than a factor of about 1000, greater than afactor of about 1500, greater than a factor of about 2000, greater thana factor of about 5000, greater than a factor of about 10,000, orgreater than a factor of about 20,000.

Procedures: Cells are thawed from a liquid nitrogen preserved state.Once cells have been expanded and divide at their expected doublingtimes, screening begins. Cells are seeded in growth media in eitherblack 1536-well or 384-well tissue culture treated plates. Cells arethen equilibrated in assay plates via centrifugation and placed inincubators attached to the Dosing Modules at 37° C. for 24 hours beforetreatment. At the time of treatment, a set of assay plates (which do notreceive treatment) are collected and ATP levels are measured by addingATPLite (Perkin Elmer). These Tzero (T₀) plates are read usingultra-sensitive luminescence on Envision plate readers (Perkin Elmer).Treated assay plates are incubated with compound for 72 hours. After 72hours, plates are developed for endpoint analysis using ATPLite. Alldata points are collected via automated processes, quality controlledand analyzed using Zalicus software. Assay plates are accepted if theypass the following quality control standards: relative luciferase valuesare consistent throughout the entire experiment, Z-factor scores aregreater than 0.6, untreated/vehicle controls behave consistently on theplate.

Growth Inhibition (GI) is used as a measure of cell viability. The cellviability of vehicle is measured at the time of dosing (T0) and after 72hours (T72). A GI reading of 0% represents no growth inhibition—T72compound-treated and T72 vehicle signals are matched. A GI reading of100% represents complete growth inhibition-T72 compound-treated and T0vehicle signals are matched. Cell numbers have not increased during thetreatment period in wells with GI 100% and may suggest a cytostaticeffect for compounds reaching a plateau at this effect level. A GIreading of 200% represents complete death of all cells in the culturewell. Compounds reaching an activity plateau of GI 200% are consideredcytotoxic. GI is calculated by applying the following test and equation:

${{If}\mspace{14mu} T} < {V_{0}\text{:}\mspace{14mu} 100*( {1 - \frac{T - V_{0}}{V_{0}}} )}$${{If}\mspace{14mu} T} \geq {V_{0}\text{:}\mspace{14mu} 100*( {1 - \frac{T - V_{0}}{V - V_{0}}} )}$where T is the signal measure for a test article, V is thevehicle-treated control measure, and V₀ is the vehicle control measureat time zero. This formula is derived from the Growth Inhibitioncalculation used in the National Cancer Institute's NCI-60high-throughput screen.

Inhibition (I) is defined asI=(1−T/V)*100%where T is treated cell count and V is untreated (vehicle) cell count(at 72 hours). I ranges from 0% (when T=V) to 100% (when T=0). The IC₅₀value is defined as the drug concentration needed to inhibit 50% of thecell growth compared to growth of the vehicle treated cells (the drugconcentration which gives I=50%). The measure of effect in theexperiment can be the inhibition of cellular response relative to theuntreated level (vehicle alone). For untreated vehicle and treatedlevels V and T, a fractional inhibition I=1−T/V is calculated. Theinhibition ranges from 0% at the untreated level to 100% when T=0.Inhibition levels are negative for agents that actually increase levels.Other effect measures, such as an activity ratio r=T/V may be moreappropriate for some assays. When activity ratios (e.g, fold increaseover stimulated control) are being used, the effect can be measuredusing an induction I=ln(T/V). With this definition, all effectexpressions are the same as for inhibition.

Combination analysis data are collected in a 6×6 dose matrix. Synergy iscalculated by comparing a combination's response to those of its singlecompound, against the drug-with-itself dose-additive reference model.Deviations from dose additivity may be assessed visually on anisobologram or numerically with a Combination Index (CI). See Table 3below for CI at 50% inhibition and CI at 50% growth inhibition. Acombination provides additive effect when CI=1.0; synergistic effectwhen CI<1.0; and antagonistic effect when CI>1.0.

Potency shifting is evaluated using an isobologram, which demonstrateshow much less drug is required in combination to achieve a desiredeffect level, when compared to the single agent doses needed to reachthat effect. The isobologram is drawn by identifying the locus ofconcentrations that correspond to crossing the indicated inhibitionlevel. This is done by finding the crossing point for each single agentconcentration in a dose matrix across the concentrations of the othersingle agent. Practically, each vertical concentration C_(Y) is heldfixed while a bisection algorithm is used to identify the horizontalconcentration C_(X) in combination with that vertical dose that givesthe chosen effect level in the response surface Z(C_(X),C_(Y)). Theseconcentrations are then connected by linear interpolation to generatethe isobologram display. For synergistic interactions, the isobologramcontour fall below the additivity threshold and approaches the origin,and an antagonistic interaction would lie above the additivitythreshold. The error bars represent the uncertainty arising from theindividual data points used to generate the isobologram. The uncertaintyfor each crossing point is estimated from the response errors usingbisection to find the concentrations where Z−σ_(Z)(C_(X),C_(Y)) andZ+σ_(Z)(C_(X),C_(Y)) cross I_(cut), where σ_(Z) is the standarddeviation of the residual error on the effect scale.

To measure combination effects in excess of Loewe additivity, a scalarmeasure to characterize the strength of synergistic interaction termedthe Synergy Score is devised. The Synergy Score is calculated as:Synergy Score=log f _(X) log f _(Y)Σ max(0,I _(data))(I _(data) −I_(Loewe))The fractional inhibition for each component agent and combination pointin the matrix is calculated relative to the median of allvehicle-treated control wells. The Synergy Score equation integrates theexperimentally-observed activity volume at each point in the matrix inexcess of a model surface numerically derived from the activity of thecomponent agents using the Loewe model for additivity. Additional termsin the Synergy Score equation (above) are used to normalize for variousdilution factors used for individual agents and to allow for comparisonof synergy scores across an entire experiment. The inclusion of positiveinhibition gating or an I_(data) multiplier removes noise near the zeroeffect level, and biases results for synergistic interactions at thatoccur at high activity levels.

The Synergy Score measure is used for the self-cross analysis. SynergyScores of self-crosses are expected to be additive by definition and,therefore, maintain a synergy score of zero. However, while someself-cross synergy scores are near zero, many are greater suggestingthat experimental noise or non-optimal curve fitting of the single agentdose responses are contributing to the slight perturbations in thescore. This strategy is cell line-centric, focusing on self-crossbehavior in each cell line versus a global review of cell line panelactivity. Additivity should maintain a synergy score of zero, andsynergy score of 2 or 3 standard deviations indicate that thecombination is synergistic at statistically significant levels of 95%and 99%, respectively.

Loewe Volume (Loewe Vol) is used to assess the overall magnitude of thecombination interaction in excess of the Loewe additivity model. LoeweVolume is particularly useful when distinguishing synergistic increasesin a phenotypic activity (positive Loewe Volume) versus synergisticantagonisms (negative Loewe Volume). When antagonisms are observed, theLoewe Volume should be assessed to examine if there is any correlationbetween antagonism and a particular drug target-activity or cellulargenotype. This model defines additivity as a non-synergistic combinationinteraction where the combination dose matrix surface should beindistinguishable from either drug crossed with itself. The calculationfor Loewe additivity is:I _(Loewe) that satisfies(X/X _(I))+(Y/Y _(I))=1where XI and YI are the single agent effective concentrations for theobserved combination effect I. For example, if 50% inhibition isachieved separately by 1 μM of drug A or 1 μM of drug B, a combinationof 0.5 μM of A and 0.5 μM of B should also inhibit by 50%.

Exemplary combinations of PI3K gamma-selective and delta-selectivecompounds were tested according to the procedures described above. Inone exemplary study, the combination effects of Compound AA withCompound BB at various concentrations were studied in various cell linesand the results are listed in Table 19 below. Compound AA is a PI3Kδinhibitor (e.g., delta-selective compound) that has a gamma/deltaselectivity ratio of greater than about 50. Compound BB is a PI3Kγinhibitor (e.g., gamma-selective compound) described herein that has adelta/gamma selectivity ratio of greater than about 50. The gamma/deltaselectivity ratio is determined by dividing the inhibitor's IC₅₀ againstPI3K gamma isoform by the inhibitor's IC50 against PI3K delta isoform.The delta/gamma selectivity ratio is determined by dividing theinhibitor's IC₅₀ against PI3K delta isoform by the inhibitor's IC50against PI3K gamma isoform. For illustrative purposes, the isobologramsof FIG. 6 (at 90% growth inhibition) and FIG. 7 (at 40% growthinhibition) show that a combination of Compound AA and Compound BBprovides synergy in diffuse large B-cell lymphoma (DLBCL) in TMD8 andFarage cell lines. Isobolograms of a combination of Compound AA andCompound BB also demonstrate synergistic effects in a B-cell lymphomacell line (Karpas-422 cell line), and in a T-cell lymphoma,non-Hodgkin's lymphoma, Hodgkin lymphoma, and anaplastic large celllymphoma cell line (HH cell line) (isobolograms not shown).

The CI₅₀ values for growth inhibition and inhibition in Table 19 arecategorized as follows: S=0.1 to <0.5, T=0.5 to <0.7, U=0.7 to <1, W=≥1.The synergy score values for growth inhibition and inhibition arecategorized as follows: A1=0.01 to <1, A2=1 to <3, and A3=>3. The typesof cell lines tested are diffuse large B-cell lymphoma (DBCL) activatedB-cell-like (ABC), DBCL germinal center B-cell-like (GCB), follicularlymphoma, mantle cell lymphoma, multiple myeloma, and T-cell lymphoma.Data show that a combination of a gamma-selective and a delta-selectivecompound provides a synergistic effect in various types of cancer celllines.

TABLE 19 Synergy CI₅₀ Score Synergy growth growth CI₅₀ Score Cell Linetype of cell line inhibition inhibition inhibition inhibition HBL-1DLBCL ABC T A2 W A1 OCT-Ly3 DLBCL ABC U A2 A1 U-2932 DLBCL ABC T A2 W A1TMD8 DLBCL ABC W A3 U A3 OCI-Ly7 DLBCL GCB W A1 A1 SU-DHL-10-epst DLBCLGCB U A3 U A2 SU-DHL-4-epst DLBCL GCB T A3 T A2 DOHH-2 DLBCL GCB S A3 SA2 Farage DLBCL GCB S A3 S A3 RL Follicular lymphoma W A1 A1 KARPAS-422Follicular lymphoma T A2 T A1 WSU-NHL Follicular lymphoma U A3 U A2GRANTA-519 Mantle cell lymphoma T A1 W A1 Jeko-1 Mantle cell lymphoma UA1 T A1 Mino Mantle cell lymphoma T A2 T A2 RPMI-8226 Multiple myeloma WA1 W A1 OPM-2 Multiple myeloma W A1 A1 NCI-H929 Multiple myeloma S A3 TA2 RH T-cell lymphoma S A2 S A1 KARPAS-299 T-cell lymphoma A1 A1

Example 253: PI3K Gamma Selective Compounds Inhibit CXCL12-InducedT-Cell Migration in Malignant B-Cell In Vitro Model

The PI3K gamma selective compounds provided herein can inhibitCXCL12-induced T-cell migration. Specifically, the material used weremedia (RPMI+0.5% Fetal Bovine Serum+Pen/Strep), total CLL PBMCs(AllCells), CoStar 24-transwell plates (5 uM insert) #3421, BD CytofixFixation Buffer (#554655), BD FBS Stain Buffer (BD Biosciences, 554656),Deep 96-well plate (Axygen, P-2ML-SQ-C), rhCXCL12 (R&D, 350-NS-050),CD-19 APC Cy7 (BD #348794 1:20), CD3-PerCPCy5.5 (BD #560835, 1:20),CD5-PE (Biolegend #300608, 1:5), CD4-FITC (BD #561842, 1:20), andCD8-APC (BD #561953, 1:20).

Total CLL human PBMCs were preincubated with Compound AA or BB in mediafor 45 minutes at 37° C. 600 uL of either basal or media containing 300ng/mL CXCL12+/−DMSO or Compound AA or Compound BB was placed in thelower chamber of transwell insert. After the 45 minutes of compoundpreincubation, 500-750K CLL PBMCs were placed in the upper chamber ofthe transwell insert in a total volume of 100 uL. The cells were allowedto migrate for 2-4 hours at 37° C. The inserts were removed, and 550 uLmedia from lower chamber were transferred into a deep 96-well plate. Thecells were spun at 1280 rpm for 5 minutes and flick media. The cellswere resuspended in 400 uL BD Cytofixation buffer and incubated at RTfor 10 minutes. 1 mL BD FBS stain buffer was added. The cells were spunat 1280 rpm and fixation buffer was disposed. The cells were resuspendedin 100 uL of CD3,4,8,5,19 antibody cocktail made up in BD FBS stainbuffer, which was then incubated at room temperature in the dark for30-60 minutes. 1 mL BD FBS stain buffer was added to each well, whichwas then spun and flicked. The cells were resuspended in 150 uL FBSstain buffer and transferred to FACS tubes already containing 150 uL FBSstain buffer. Each sample was read on the FACS for 25 seconds. Gate onCD3, 4, 8 and CD19/5 subpopulations individually and calculate averagemigration index from triplicate samples. See e.g., Borge et al.,haematologica 2010, 95(5): 768-775; de Rooij et al., Blood 2012, 119:2590-2594.

Compound AA is a PI3K delta selective compound described herein that hasa gamma/delta selectivity ratio of greater than about 50. Compound BB isa PI3K gamma selective compound described herein that has a delta/gammaselectivity ratio of greater than about 50. FIG. 8 shows the percentcell inhibition of CXCL12-induced CD3+ T cell migration at variousconcentrations of Compound AA (EC₅₀=694 nM) and Compound BB (EC₅₀=10nM). In a similar experiment, the EC₅₀ was determined for three subsetsof T-cells for a PI3K-δ inhibitor (Compound AA) and a PI3K-γ inhibitor(Compound BB). The results are shown in the following Table. Compound BBis a potent inhibitor of CXCL12 induced T-cell migration.

TABLE 20 AVG Cmpd AA Cmpd BB T-cell subset EC₅₀ (nM) EC₅₀ (nM) CD3⁺ 630± 71  17 ± 17 CD4⁺ 726 ± 230 20 ± 21 CD8⁺ 423 ± 290 13 ± 15

The data demonstrate that Compound BB (a gamma selective compound) ismore potent than Compound AA (a delta selective compound) at inhibitingCXCL12-induced CD3+ T cell migration in CLL PBMCs. Gamma selectivecompounds can be used to block the migration of growth promoting T-cellsinto the B-cell tumor niche slowing down the progression of the disease.This could translate to increased progression free survival or deeperresponses in the clinic with respect to B cell malignancies.

The mechanism of Compound BB's inhibition of migration was studied. Itwas found that Compound BB inhibited CXCL12-induced pAKT in T-cells morestrongly than a PI3K-δ inhibitor (Compound AA) (FIG. 9). There was atight correlation between the EC₅₀s observed in the CXCL12-induced pAKTassay and the T-cell migration assay. This result suggests that a PI3K-γinhibitor interferes with T cell migration in CLL patient PBMCs byblocking pAKT signaling. While PI3K inhibition does not abrogateIL4-induced CLL survival (data not shown), PI3K-γ-dependent inhibitionof T-cell migration to CLL lymph node microenvironments may indirectlyprevent T-cell derived cytokines (e.g., IL-4) from providing PI3Kindependent CLL survival signals.

Without being bound by a particular theory, the data suggest thefollowing model in B cell malignancies. Malignant B-cells receive growthand pro-survival signals from BCR activation and mesenchymal cells,transmitted via the PI3K-δ, and-γ pathways. Malignant B-cell growth andsurvival is also supported by T-cells and myeloid cells viaPI3K-dependent and -independent mechanisms. Inhibition of PI3K-δ reducesthe proliferation of malignant B-cells; however, other PI3K-independentsignals may potentiate tumor cell survival. Inhibition of PI3K-γ, e.g.,by Compound BB, may block migration and/or differentiation of accessoryT-cells and myeloid cells and abolish these key support cells within thetumor microenvironment. The lack of key support cells within the tumormicroenvironment as a result of PI3K-γ inhibition may thus morecompletely block malignant B-cell growth and survival. Without beingbound by a particular theory, Compound BB may act to inhibit tumorgrowth and/or survival at any of the above-mentioned steps.

Example 254: Inflammatory Bowel Disease (IBD) Model

A study can be conducted to evaluate the potential efficacy of acompound provided herein in female SCID mice with CD4⁺ inflammatorybowel disease. In this murine model, female C.B-17 SCID mice areinjected intraperitoneally (IP) with CD45RB^(high) cells, a subset ofCD4⁺ T cells obtained from normal BALB/c mice, to induce spontaneouschronic inflammation in the large intestine. Gross and histopathologicchanges resulting from this treatment resemble those occurring inCrohn's disease and ulcerative colitis in humans. See Leach et al.,Inflammatory Bowel Disease in C.B-17 scid Mice Reconstituted with theCD45RB^(high) Subset of CD4⁺ T Cells, American Journal of Pathology,1996, 148 (5), 1503-1515.

On study day 0, Balb/C mice are terminated, and spleens are obtained forCD45RB^(high) cell isolation per the SCID IBD cell separation protocol.SCID mice are weighed and received intraperitoneal (IP) injections ofthe sorted cells (approx. 4×10⁶ cells/ml, 100 μl/mouse injections). Onstudy day 21, mice are weighed and randomized by body weight loss intotreatment groups, and daily (QD), oral (PO) dosing is initiated. Dosingcontinued through study day 41, and mice are terminated on day 42. Themice can be divided into various groups. Group 1 can be the normalcontrol; group 2 can be the vehicle control (0.5% CMC, 0.05% Tween80 inH₂O); and various amounts of a compound provided herein can beadministered to different groups as comparison.

For each animal, the entire colon (proximal and distal) is trimmed into8 equally spaced pieces for processing and embedding Sections arestained with hemotoxylin and eosin (H&E). For each H&E stained section,submucosal edema is quantitated by measuring the distance from themuscularis mucosa to the internal border of the outer muscle layer in anon tangential area thought to most represent the severity of thischange. Mucosal thickness is also measured in a non-tangential area ofthe section that best represented the overall mucosal thickness. Thisparameter is indicative of gland elongation and mucosal hyperplasia. Inorder to incorporate this parameter into the summed score, a hyperplasiascore is derived from the measurement as follows: 0=<250 μm; 1=250-349μm; 2=350-449 μm; 3=450-599 μm; 4=600-699 μm; and 5=≥700 μm.

The extent of inflammation (foamy macrophage, lymphocyte and PMNinfiltrate) is assigned severity scores according to the followingcriteria:

Normal=0

Minimal=1 (generally focal affecting 1-10% of mucosa or if diffuse thenminimal)

Mild=2 (generally focal affecting 11-25% of mucosa or if diffuse thenmild)

Moderate=3 (26-50% of mucosa affected with areas of gland loss replacedby inflammatory cell infiltrate, milder in remaining areas of mucosa)

Marked=4 (51-75% of mucosa affected with areas of gland loss replaced byinflammatory cell infiltrate, milder in remaining areas of mucosa)

Severe=5 (76-100% of mucosa affected with areas of gland loss replacedby inflammatory cell infiltrate, milder in remaining areas of mucosa)

The parameters reflecting epithelial cell loss/damage are scoredindividually using a percent area involved scoring method: None=0; 1-10%of the mucosa affected=1; 11-25% of the mucosa affected=2; 26-50% of themucosa affected=3; 51-75% of the mucosa affected=4; and 76-100% of themucosa affected=5.

Parameters that are scored using percent involvement included: (1) Colonglandular epithelial loss—this includes crypt epithelial as well asremaining gland epithelial loss; and (2) Colon Erosion—this reflectsloss of surface epithelium and generally is associated with mucosalhemorrhage (reflective of the bleeding seen clinically and at necropsy).

The 4 important scored parameters (inflammation, glandular epithelialloss, erosion, hyperplasia) are ultimately summed to arrive at a sum ofhistopathology scores, which indicates the overall damage and would havea maximum score of 20.

Inflammatory cell infiltrates in the colonic mucosa are evaluated forapproximate % of neutrophils in the total infiltrate using the criteriabelow. The approximate % of total is then multiplied by the 0-5inflammation score in an attempt to semiquantify relative PMNinfiltration across sections and animals: 0=approx 0%; 10=approx 10%;25=approx 25%; 50=approx 50%; and 75=75% or greater. This value is thenmultiplied by the inflammation score in an attempt to semiquantifyrelative PMN infiltration across sections and animals.

Example 255: Autoimmune Encephalomyelitis (EAE) Model

Effects of a compound provided herein on treating inflammation can betested in a 28 day semi-therapeutic mouse EAE model. EAE can be inducedin 60 mice using Hooke Kit™ MOG₃₅₋₅₅/CFA Emulsion PTX (HookeLaboratories, Lawrence Mass.), catalog number EK-0110, lot number 0104,per manufacturer's recommended protocol.

Mice are injected subcutaneously, at two sites in the back, with theemulsion component of the kit (containing MOG₃₅₋₅₅). One site ofinjection is in the area of the upper back, approximately 1 cm caudal ofthe neck line. The second site is in the area of the lower back,approximately 2 cm cranial of the base of the tail. Injection volume is0.1 mL at each site. Within 2 hours of the injection of emulsion, andthen again 24 hours after the injection of emulsion, the pertussis toxincomponent of the kit is administered intraperitoneally. Volume of eachinjection is 0.1 mL.

EAE is induced in certain mices, which can be divided into variousgroups e.g., a group of negative control and groups treated with variousamounts of a compound provided herein.

Readouts are EAE scores and changes in body weight. Mice are scoreddaily. EAE is scored on scale 0 to 5:

0—No obvious changes in motor functions of the mouse in comparison tonon-immunized mice. When picked up by the tail, the tail has tension andis erect. Hind legs are usually spread apart. When the mouse is walking,there is no gait or head tilting.

1—Limp tail. When the mouse is picked up by the tail, instead of beingerect, the whole tail drapes over finger.

2—Limp tail and weakness of hind legs. When mouse is picked up by tail,legs are not spread apart, but held closer together. When the mouse isobserved walking, it has a clearly apparent wobbly walk.

3—Limp tail and complete paralysis of hind legs (most common); or limptail with paralysis of one front and one hind leg; or all of: severehead tilting, walking only along the edges of the cage, pushing againstthe cage wall, and spinning when picked up by the tail.4—Limp tail, complete hind leg and partial front leg paralysis. Mouse isminimally moving around the cage but appears alert and feeding. Usually,euthanasia is recommended after the mouse scores level 4 for 2 days.When the mouse is euthanized because of severe paralysis, score of 5 isentered for that mouse for the rest of the experiment.5—Complete hind and complete front leg paralysis, no movement around thecage; or mouse is spontaneously rolling in the cage; or mouse is founddead due to paralysis.

At the end of the study, the following tissues can be collected andanalyzed, e.g., collection of plasma for PK, collection of serum,collection of spines for histological analysis, histological analysis ofspines, count of inflammatory foci, estimation of demyelination area, orcount of apoptotic cells.

Example 256: Pharmacokinetics and Reduction of LPS-InducedNeutrophilia—Oral Administration

To evaluate the effect of Compound BB (PI3Kγ inhibitor (e.g.,gamma-selective compound provided herein)) and Compound AA (PI3Kδinhibitor (e.g., delta-selective compound)) on the LPS inducedinflammatory parameters, Compound BB and Compound AA were administeredorally. Compound AA has a gamma/delta selectivity ratio of greater thanabout 50. Compound BB has a delta/gamma selectivity ratio of greaterthan about 50.

This study also includes assessment of the exposure to the compounds inplasma and lung tissues. The groups and details of the administrationare provided in the table below.

TABLE 21 LPS delivered intra-tracheally (i.t.) Number Groups (mg/kg) ofrats 1. vehicle/saline i.t. volume 0.25 mL/rat 6 2. vehicle/LPS i.t. 1ug LPS/rat 10 3. Compound AA (p.o.) 0.2 mg/kg 7 4. Compound AA (p.o.) 1mg/kg 7 5. Compound AA (p.o.) 5 mg/kg 7 6. Compound BB (p.o.) 0.5 mg/kg7 7. Compound BB (p.o.) 1.5 mg/kg 7 8. Compound BB (p.o.) 5 mg/kg 7 9.Ref A (p.o.) 3 mg/kg 7 Reference A = budesonide.p.o. vehicle for Compound AA and Compound BB is 5% NMP and 95% PEG 400.Ref C is budesonide p.o. The compound was administered p.o. and about 1hour later, LPS was administered intra-tracheally and plasma wascollected. After about 5 hours, plasma and bronchoalveolar lavagesamples were collected.

The oral PK summary of Compound AA and Compound BB in the rat LPSstudies is provided in the table below.

TABLE 22 Free Plasma Cone (nM) Cellular IC₅₀ (nM) Dose (mg/kg) 1.5 h*5.5 h** δ γ β α Compound 0.5 2.1 2.6 197 1 189 873 BB 1.5 5.2 5.4 5 22.624.7 Compound 0.2 BLQ BLQ 0.1 418 102 1900 AA 1 1.8 1.1 5 17.8 4.6 * =free plasma concentration at time of LPS challenge. ** = free plasmaconcentration approximately 4 hours post LPS challenge.

Compound BB and Compound AA (p.o.) reduced neutrophil in bronchoalveolarlavage fluid (BALF) 4 hours post-LPS challenge in a dose dependentmanner. Compound BB had a more profound effect on neutrophil influxcompared Compound AA (FIG. 10, FIG. 11 and FIG. 12 (mean with SEM,p-value is from t-test; groups are individually compared to theLPS-group)). These data show that PI3K-γ is involved in infiltration ofneutrophils into the lung, a key pathologic cell type in chronicobstructive pulmonary disease (COPD).

PK results showed free plasma levels of both Compound BB and Compound AAat the highest dose levels were equal or greater than the IC₉₀ for gammaand delta, respectively, and yet well below the IC₅₀ for the nextclosest PI3K isoforms for each compound.

In another exemplary study, it was found out that the increase inCXCL12-induced p-AKT in CD3+ T cells was mediated by PI3K-γ. In yetanother exemplary study, it was found out that, within the malignantB-cell population, the increase in CXCL12-induced p-AKT was PI3K-δdependent, suggesting that CXCL12 signals through different PI3Kisoforms in these varying cell types.

Example 257: Effects on CXCL12 Induced p-AKT Induction in DifferentiatedMacrophages

Myeloid-derived cells and mesenchymal stromal cells can support CLL cellsurvival as components of the tumor microenvironment (TME). Recentreports suggest that CLL cytoprotective nurse-like cells may have an M2polarization and be similar to the immunosuppressive tumor associatedmyeloid cells found in solid tumors. Giannoni et al., Haematologica2014, 99(6), 1078-88. A purpose of this study is to model these TMEcomponents and to evaluate the effects of PI3K isoform selectiveinhibitors on CXCL12 induced p-AKT induction in differentiatedmacrophages. Compound AA is a PI3K delta selective compound that has agamma/delta selectivity ratio of greater than about 50. Compound BB is aPI3K gamma selective compound described herein that has a delta/gammaselectivity ratio of greater than about 50.

Macrophage Differentiation and Polarization:

Long bones from young C57B6 mice were harvested and bone marrow wasextruded by gentle lavage of the bone marrow cavities with a 28 gaugeneedle. Red blood cells were removed using red blood cell lysis buffer(Sigma). Bone marrow cells were differentiated to macrophages using MCSFfor 6 days (RPMI and 20% FCS) with one media change (with removal of allnon-adherent cells) at day 3. Macrophages were polarized into M0 (MCSF),M1 (MCSF, IFNγ, and LPS for 1 day), or M2 (MCSF, IL4 for 2 days).

CXCL12-Induced p-AKT Induction Assay:

Polarized macrophages were lifted with Acutase, and plated at 0.5M cellsper well (0.5 ml) in a deep well (2 ml) non adherant plate inpolarization media for 1 hour. DMSO or tested compound in DMSO (0.1%final DMSO concentration, Compound AA or Compound BB) was added for 40minutes. CXCL12 was added, and the cells were mixed on a plate shakerfor 2 minutes. 1 ml of 37 degree FACS Lyse/fix buffer (BD bioscience)was added and the plate was mixed for 1 minute. The plates were spun anddecanted and frozen at −80° C. Phospho AKT staining with anti PhosphoAKT 473 PERCP labeled (Cell Signaling Technologies) was performed andmeasured by flow cytometry. Total cell MFI was used to calculate theeffect of tested compound on CXCL12 induction of phospho AKT.

Results:

Compound BB inhibited CXCL12 induced p-AKT in differentiated and M1polarized macrophage with an IC₅₀ value of about 14 nM. Compound BBinhibited CXCL12 induced p-AKT in differentiated and M2 polarizedmacrophage with an IC₅₀ value of about 8.4 nM. Compound AA did notsignificantly inhibit CXCL12-induced p-AKT in differentiated and M1 orM2 polarized macrophages. FIG. 13 shows the p-AKT staining result for M2phenotype. The results of this study show that CXCL12 mediated-M2activation is dependent upon PI3K-γ as it is more potently inhibited bythe PI3K-γ selective inhibitor (Compound BB) than the PI3K-δ selectiveinhibitor (Compound AA).

Example 258: CLL Co-Culture with M2 Macrophage Cells

Frozen purified CLL cells were obtained from a commercial vendor (AllCells). Macrophages were polarized into M0 (MCSF) or M2 (MCSF, IL4 for 2days). M0 or M2 polarized macrophages were plated into a 96 well tissueculture plate at the stated density in RPMI+10% FCS with theirrespective polarization factors. The CLL cells were plated in the tissueculture plate at 150 thousand cells per well with media or M0 or M2macrophages. CLL cell viability over time was measured by Guava viacount(Millipore). Co-cultures of M2 macrophages with CLL cells led toextended CLL cell survival (FIG. 14). The result shows that M2macrophage cells have an ability to protect CLL cells from cell death,that is enhanced over the protective effects of the M0 cells at 120hours (data not shown).

The results indicate that PI3K-γ selective compounds can be used totreat CLL by potently inhibiting M2 activation and reducing survival ofCLL cells. FIG. 15 is a schematic of the differentiation of myeloidprogenitor cells and interactions between certain T cells. FIG. 16illustrates differentiation of a myeloid cell into an M1 macrophage orM2 macrophage.

Example 259: Effect on Macrophage Differentiation into Osteoclasts

Recent reports suggest that PI3K-γ knockout mice have normal osteoblastactivity, but are deficient in osteoclast activity, which leads to anincrease in bone formation. Osteoclast differentiation from bone marrowcells is deficient in these PI3K-γ knockout mice. Kang et al., Proc NatlAcad Sci USA. 2010, 107(29): 12901-12906. A purpose of this study is toevaluate PI3K-γ selective inhibitor on differentiation of osteoclastsfrom bone marrow macrophages. Compound BB is a PI3K gamma selectivecompound described herein that has a delta/gamma selectivity ratio ofgreater than about 50.

Bone Marrow Derived Macrophages as Osteoclast Progenitors:

Long bones from young C57B6 mice were harvested and bone marrow wasextruded by gentle lavage of the bone marrow cavities with a 28 gageneedle. The cells were screened, and red blood cells were removed usingred blood cell lysis buffer (Sigma). Bone marrow cells weredifferentiated to macrophages using 20 ng/ml mouse MCSF (RnD systems)for 3 days (RPMI and 20% FCS). The cell were washed with PBS, and liftedwith Accutase for 10 minutes.

Osteoclast Differentiation Assay:

The bone marrow macrophage cells were plated at 150,000 cells per wellin a 96 well plate in a MEM 10% FCS. The cells were treated with DMSO orCompound BB in DMSO for 40 minutes prior to cytokine addition. Then 25ng/ml mouse MCSF with or without mouse RANKL (100 ng/ml, RnD systems)was added for 6 days. Media changes (including drugs) were done at day 3and day 5. The cells were then fixed and stained with the tartrateresistant acid phosphatase (TRAP) assay kit from Sigma according to themanufactures instructions. TRAP positive cells (osteoclasts and cellsundergoing an osteoclastic differentiation) in a transect of thetriplicate wells (at 20× concentration) were manually counted by goingthrough the widest part of each well.

Results:

The results are depicted in FIG. 17, which shows that Compound BBprevents RANKL driven differentiation of osteoclasts from bone marrowmacrophages. The results indicate that PI3K-γ selective compounds can beused for treatment of osteoclast driven bone diseases.

Example 260: Pharmacokinetics

The purpose of the study is to evaluate the pharmacokinetics of CompoundBB following a single and a repeat dose administration in CB6F1 femalemice. Compound BB was orally administered at 2, 7.5 and 15 mg/kg oncedaily for 3 consecutive days to female mice (Female CB6F1 mice, n=72(5-6 weeks old). The animals were divided into three dosing groups:Compound BB @ 2 mg/kg, PO in 5% NMP, 95% PEG400; Compound BB @ 7.5mg/kg, PO in 5% NMP, 95% PEG400; and Compound BB @ 15 mg/kg, PO in 5%NMP, 95% PEG400.

Compound BB was administered orally once a day for a total of three daysat a dosing volume of 10 mL/kg based on the most recent body weight.Animals were weighed daily prior to dosing. On the day of the first doseanimals were closely monitored over the 1st hour followingadministration, animals were observed at least twice during the 4 hoursfollowing dose administration, and at the end of the day. On subsequentdosing days, the animals' condition was noted prior to dosing; theanimals were observed within 4 hours of dosing, and at the end of theday. Increased monitoring can occur if the animals are exhibitingclinical signs. If an animal is exhibiting mild to moderate clinicalsigns of toxicity, dosing may be held, in consultation with the studydirector, to allow for recovery.

Samples were collected at pre-dose 0.25, 0.5, 1, 2, 4, 6 and 24 hfollowing the first dose on day 1 and pre-dose (trough), 0.25, 0.5, 1,2, 4, 6 and 24 h following the final dose on day 3. Animals received asingle oral dose daily for 3 consecutive days. Blood was collected byretro-orbital bleeding for all time points on day 1. All other bloodsampling was collected by a cardiac puncture. Blood was collected intoK2EDTA tubes and placed on wet ice. Within 15 min of collection, thesample was centrifuged at 10,000 rpm at 4° C. for 10 min in a bench topcentrifuge and the plasma transferred to a new microcentrifuge tube.

FIG. 18 shows the mean unbound concentration of Compound BB in plasma, 1or 5 hours after dosing. FIG. 19 shows the results of the study above,for measuring a 24-hour time course of unbound plasma concentration ofCompound BB. This experiment indicates that Compound BB has goodstability in serum and is suitable for administration, e.g., once perday.

A similar study was carried out using Compound 88. FIG. 20 shows themean plasma unbound Compound 88 concentration-time profiles for femaleC57 mice administered 2, 7.5 or 15 mg/kg PO. Below is a table thatdetails the results for Compound 88:

C_(max) AUC_(last) AUC₀_INF AUC₀₋₈ (ng/ t_(max) t_(last) (ng*h/ (ng*h/(ng*h/ t_(1/2) mL) (h) (h) mL) mL) mL) (h)   2 mg/kg 654 2 8 3891 N/A3891 N/A PO NMP 7.5 mg/kg 3705 1 24 20625 20684 15819 2.8 PO NMP  15mg/kg 5705 1 24 71746 81234 34591 7.9 PO NMP N/A = Not applicable due toinsufficient data points in terminal phaseThis shows that Compound 88 also has good stability in serum and issuitable for administration.

Example 261: Effects of Compound BB on T Cell Activation

The purpose of this study is to evaluate the effect of various antigens(Con-A) in T cell activation in Human Whole Blood vs. 1:1 Diluted Bloodusing test compounds at 24 hour time-point.

T-cell Activation Media includes RPMI-1000 ml, MEM NEAA-10 ml, SodiumPyruvate (100 mM)-10 ml, Pen Strep-10 ml, 2-Mercaptoethanol-1 ml, andFBS-100 ml. 100 ul of whole blood were plated in 96 well plate. Usedonly the 60 inside wells and filled the outside with media or PBS.Diluted human whole blood with 5% Serum RPMI 1640 1:1 and plate 100 ulin 96 well plate. Used only the 60 inside wells and filled the outsidewith media or PBS. The samples were allowed to rest for 45 minutes inthe incubator, and then they were treated with Compound BB for 45minutes.

The compound was diluted using DMSO (Sigma hybridmax). For 1000 nMhighest concentration starting from a 10 mM stock, added 300 ul of DMSOinto deep well plates. To the highest concentration add 10 ul of 10 mMcompound stock. See table below.

DMSO DMSO DMSO DMSO DMSO DMSO DMSO DMSO (200 ul) (200 ul) (200 ul) (200ul) (200 ul) (200 ul) (200 ul) (200 ul) DMSO (50 ul (50 ul (50 ul (50 ul(50 ul (50 ul (50 ul (50 ul (200 ul) + from from from from from fromfrom from DMSO Compound previous previous previous previous previousprevious previous previous (200 ul) stock well) well) well) well) well)well) well) well) compound 10 ul

A media plate was prepared from DMSO plate by taking 100 ul and go into900 ul media in new deep well plate, taping plate, and putting on mixerfor 5 minutes.

The rested cell plates were removed from incubator. Added 12 uL ofdiluted compound from media plate (final DMSO is 1%), and incubated for45 minutes.

Con-A was prepared as follows: 200 ul stock into 1 ml Media, 100 ul ofthat into 1 ml Media, 100 ul of that into 1 ml Media. Used highestconcentration. This was analyzed after 24 Hours.

10× stock in media, added 12 ul per well. Incubated at 37° C. degree inhumidity chamber for 24 hours at 5% CO₂. Analyzed with multiplex elisafrom MSD. Human Pro-Inflammatory Panel 1 IFNg V-Plex Kit.

Media CON-A CON-A CON-A CON-A CON-A CON-A CON-A CON-A CON-A Media CON-ACON-A CON-A CON-A CON-A CON-A CON-A CON-A CON-A Media CON-A CON-A CON-ACON-A CON-A CON-A CON-A CON-A CON-A CON-A CON-A CON-A CON-A CON-A CON-ACON-A CON-A CON-A CON-A CON-A CON-A CON-A CON-A CON-A CON-A CON-A CON-ACON-A CON-A CON-A CON-A CON-A CON-A CON-A CON-A CON-A CON-A CON-A CON-A

FIG. 21 shows the effects of Compound BB and Compound AA on T cellactivation, as measured by inhibition of IFN-γ. Since compound BB ishighly selective for PI3K-γ and Compound AA is highly selective forPI3K-δ, the compounds' very different IC₅₀s in this assay (3 nM forCompound AA and 2500 nM for Compound BB) indicate that PI3K-δ isimportant for T cell activation, and PI3K-γ less so.

Example 262: Effects of Compound BB on Murine M2 TAM Differentiation

Compound BB, which selectively inhibits PI3K-γ, and Compound AA, whichis highly selective for PI3K-δ, were tested for their ability to blockdifferentiation of myeloid cells into M2 TAMs, and the results are shownin FIG. 22A and FIG. 22B, respectively. High ARG1 (Arginase-1) levelsindicate that myeloid cells are differentiating into M2 macrophages, solower ARG1 levels relative to a control indicate interference withdifferentiation into M2 macrophages. Expression of Arginase-1 depletesthe tumor microenvironment of arginine, thereby promoting T cell deathand NK cell inhibition. Schmid et. al., Proceedings: AACR 103rd AnnualMeeting 2012, Cancer Research: Apr. 15, 2012; Volume 72, Issue 8,Supplement 1. Compound BB inhibited M2 TAM differentiation much moreeffectively than PI3K-δ inhibitor. M2 TAMs promote cancer cell survival(FIG. 23). Because Compound BB prevents differentiation to pro-tumor M2cells, this experiment indicates that Compound BB is expected toincrease a subject's anti-tumor immune response.

A similar experiment was performed on bone marrow cells differentiatinginto MDSCs. Compound BB suppressed both VEGF (FIG. 24A) and ARG1 (FIG.24B) in an Myeloid Derived Suppressor Cell differentiation assay.

A similar experiment was performed on M1 differentiation. Compound BBdid not have an observable effect on M1 macrophage differentiation (datanot shown). Accordingly, Compound BB is not expected to interfere withthe anti-tumor activities of M1 macrophages.

Example 263: Single Agent Activity of Compound BB in the Lewis LungCarcinoma Model

Compound BB was tested in the Lewis Lung Carcinoma model, which is knownin the art and briefly described hereinabove.

LLC tumor brei model was prepared and injected subcutaneously into amouse in the hind flank region. The tumor brei was prepared as follows.LL2-Luc tumor cells were grown in culture in DMEM+10% FBS. Cells wereimplanted @ 1×10⁶ cells/100 uL SC in the flank of C57 Albino mice (male)When tumors reached 1000 mm3, tumors were harvested. Only the white,living tissue was harvested. Any material containing visable blood ornecrotic area was isolated away from the tumor tissue. Tumor tissue wasplaced in a Dounce tissue homoginized with 5 mLs of warm DMEM (no FBS).About 1-2 grams of tumor tissue was collected, homogenized, pooled into50 mL falcon tube, and cells were counted. Cells were spinned @ 12,000RPM, and FBS was pooled off. Cells were resuspended to equal 1×10⁷cells/mL with sterile PBS. 100 uL of cells/mouse SC at the hind flankregion were implanted. This process was repeated one more time beforeconducting brei tumor efficacy study.

FIGS. 25A and 25B show that Compound BB demonstrated single agentactivity in the Lewis Lung Carcinoma model. FIG. 26 shows the meanunbound Compound BB concentrations in tumor and plasma following 12 daysof daily oral administration in NMP solution. Below is a table ofdetails:

Dose Time Free Conc SD Free Tissue Group (h) (nM) Conc Plasma   3 mg/kgTrough 0.75 0.408 2 h 25.9 14.0 7.5 mg/kg Trough 8.99 7.21 2 h 99.0 66.3 15 mg/kg Trough 8.39 3.67 2 h 157 72.2The results indicate that exposure increased with increase in dose. At 2h, Compound BB levels are above cellular gamma IC₅₀ in all groups. For7.5 mg/kg and 15 mg/kg groups, they are above the gamma IC₅₀ at 24 h.

Example 264: Effects of Compound BB on Tumor Growth in CT26 and MC38Colon Cancer Models

The purpose of this study is to determine the efficacy of Compound BB asa single agent and in combination with anti-PD-L1 in the CT26 allograftmodel of mouse colon cancer. CT26 is anN-nitroso-N-methylurethane-(NNMU) induced, undifferentiated coloncarcinoma cell line. It was cloned to generate the cell line designatedCT26.WT (ATCC catalog number CRL-2638). 5-6 week old female, Balb/c mice(Jackson Labs) were implanted subcutaneously with 5×10⁵ CT26 cells in200 μl PBS. CT26 cells were cultured in DMEM supplemented with 10% FBS.Dosing commenced when tumors reached an average volume of −100 mm³.Administration of the compounds was performed according to the followingTable:

TABLE 23 Cpnd BB α-PD-L1 PO IP QD Q3D Group # mice/group (n) mg/kgμg/mouse 1 15 0 0 2 15 6 0 3 15 0 200 4 15 6 200

Compound BB vehicle: 5% NMP/95% PEG400

Compound BB prepared at 0.6 mg/ml in 5% NMP/95% PEG400

Isotype control (Ultra-LEAF purified rat IgG2b, k isotype control, cloneRTK4530, Lot B180477, Biolegend, Cat #70640)

Anti-PD-L1 antibody (Ultra-LEAF purified anti-mouse CD274-B7H1, PD-L1-,clone 10F9G2, Lot B178331, Biolegend, Cat #124318)

Dosing Regimen:

Compound BB: QD for 3 weeks at a volume of 5 ml/kg

Anti-PD-L1: every third day for a total of 4 doses

Body weights and tumor volumes were measured three times per week.

FIG. 27A and FIG. 27B show Compound BB administered together with ananti-PD-L1 checkpoint inhibitor in the CT26 model. FIG. 27A shows theeffect on tumor volume over time. FIG. 27B shows the percent survivalover time.

FIG. 28 shows that MDSC expansion after cyclophosphamide treatmentoccurs within the 5-9 day time frame and that this expansion isdecreased by Compound BB in the CT26 model.

A second model for colon cancer, the M38 model, was used to furtherevaluate the effects of Compound BB. In this model, a syngeneic coloncancer cell line is run subcutaneously. The experimental setup was asfollows.

Female CR C57BL/6 mice were set up with 1×106 MC38 tumor cells in 0%Matrigel sc in flank. The cell injection volume was 0.05 ml per mouse.When tumors reached an average size of 80-100 mm³, a pair match was madeand treatment began. The tumor was measured by caliper biweekly.

The dosing and formulations were as follows: 5-FU in D5W; Compound BB in5% NMP/95% PEG. The dosing volume was 5 mL/kg.

For all animals, blood was collected by terminal cardiac puncture underisoflurane anesthesia. The blood was processed for plasma. In addition,tumor samples were collected.

FIG. 29A shows that Compound BB monotherapy significantly inhibitedtumor growth in the MC38 colon cancer model. FIG. 29B shows the effectsof Compound BB and 5-FU used separately and in combination in the MC38model.

At study end, tumors were isolated, fixed in 10% neutral bufferedformalin and embedded in paraffin for histologic analysis. Tumorsections from both vehicle and Compound BB monotherapy were cut forimmunohistochemical analysis. T-cells within the tumor sections wereidentified utilizing a CD3-specific antibody via automatedimmunohistochemistry equipment (Ventana). Once stained, slides weredigitalized via Aperio slide scanning equipment. Next, stained tumorsections were quantified via Definians software to assess the percentageof CD3 positive cells within each tumor. FIG. 30 shows that MC38 tumorstreated with Compound BB have significantly more intratumoral CD3+ Tcells when compared to vehicle. Final tumor volume was plotted as afunction of percent CD3+ T-cells invehicle sample (FIG. 31A) andCompound BB monotherapy sample (FIG. 31B), respectively. A trendline wasadded to the vehicle and Compound BB montherapy graphs. The vehiclefinal tumor volumes did not correlate with percent CD3+ T-cells whereasin the cohort of Compound BB monotherapy final tumor volumes stronglycorrelated with intratumoral percent CD3+ T-cells (R2=0.7873). Theresults indicate that significant influx of CD3+ T-cells upon PI3K-gammainhibition leads to decreases in tumor size.

Example 265: Effect of Compound BB in the DoHH2 Human Follicular B CellLymphoma Model

The purpose of this study is to evaluate the anti-tumor activity ofCompound AA and Compound BB alone and in combination in the DoHH2 humantransformed follicular B cell lymphoma subcutaneous model in femaleCB17.SCID mice. DoHH2, a human follicular B cell lymphoma cell line, wasgrown in tissue culture (filtered RPMI-1640 supplemented with 10% fetalbovine serum) and implanted (5×10⁶ cells in 100 uL RPMI-1640 media and100 uL matrigel (LDV free)) subcutaneously into the hind flank of femaleCB17.SCID mice. Treatment began when tumors reached 100 mm³ in volume.In dosing groups of 15 mice per group, drug was administered asfollows: 1. Vehicle (5% NMP 95% PEG400)+Vehicle; 2. Compound AA (10mg/kg)+Vehicle; 3. Compound BB (15 mg/kg)+Vehicle; 4. CompoundAA+Compound BB. Animals were dosed for 21 consecutive days. Efficacycomparisons were determined by tumor caliper measurements, three timesper week.

FIGS. 32A and 32B show the result of inhibiting PI3K-γ (Compound BB)and/or PI3K-δ (Compound AA) in the DoHH2 model. FIG. 32B is a repeatstudy. Dual inhibition of PI3K-δ and PI3K-γ is superior to inhibition ofeither isoform alone.

Example 266: Effects of Compound BB and/or PDL-1 Inhibition on TumorGrowth in Orthotopic 4T1 Breast Cancer Model

The PI3K-γ selective inhibitor, Compound BB, and the immune checkpointtherapy, PDL-1, were assayed separately and together for effects ontumor growth in the 4T1 breast cancer model. This model system isdescribed in more detail in Example 227. Briefly, in this study,treatment was initiated when tumors reached approximately 100 mm³.Compound BB or vehicle was orally administered once daily for 3 weeks.Anti-PD-L1 or isotype control antibodies were adminsteredintraperitoneal once every 3 days for a total of 5 doses. In a sample of15 mice, dosing groups were as follows. 1. Vehicle+Isotype control (RatIgG2b x, 200 ug); 2. Compound BB (7.5 mg/kg), PO+Isotype control (200ug); 3. Vehicle+anti-PD-L1 (200 ug); 4. Compound BB (7.5 mg/kg),PO+anti-PD-L1 (200 ug). During the experiment, tumor luciferin flux wasmeasured twice a week. Efficacy comparisons were determined by tumorcaliper measurements.

FIG. 33A shows reduced tumor growth with Compound BB and/or PDL-1inhibition in orthotopic 4T1 breast cancer model. FIG. 33B shows reducedtotal luciferase flux with Compound BB and/or PDL-1 inhibition inorthotopic 4T1 breast cancer model. FIG. 34 shows that Compound BBdemonstrated single agent activity in subcutaneous 4T1 breast cancermodel and that the anti-tumor effect of Compound BB was maintained forat least 6 days after the discontinuation of treatment with Compound BB.

The 4T1 model was also used to determine the anti-cancer activity ofCompound BB and 5-FU, docetaxel, or paclitaxel. Briefly, treatment beganwhen tumors reached ˜50 mm³ in volume. Compound BB or vehicle was orallyadministered once daily at 15 mg/kg in the NMP formulation to balb/cfemale mice for 2 weeks. Docetaxel 33 mg/kg Q7D×2, paclitaxel 10 mg/kgQ5D×3, 5-FU 50 mg/kg QOD×7 or saline was adminstered intraperitoneal.Efficacy was determined by tumor volume and luciferin measurements.Tumor measurements and body weights were taken three times a week.Luciferin measurements were taken twice a week.

FIG. 35 shows that Compound BB alone achieved a reduction in tumorgrowth, while co-administration of Compound BB and 5-FU achieved afurther reduction. FIG. 36 shows that Compound BB demonstrated singleagent activity in subcutaneous 4T1 breast cancer model, whileco-administration with docetaxel, paclitaxel, or 5-FU achieved a furtherreduction in tumor growth.

In some cases, a tumor being treated with an immune therapy may undergoa transient increase in size, see 2008 ASCO Abstract #3120 Wolchok.However, this increase in size is not necessarily due to cancer cellgrowth. Instead, it can be due to tumor infiltration by anti-tumorlymphocytes.

To summarize, Compound BB showed activity in models for breast cancer,lung cancer, colon cancer, and glioblastoma.

Example 267: Pharmacodynamic Studies in Murine Syngeneic Breast, Lung,Colon Models

The effect of Compound BB on solid tumor immune infiltrates wascharacterized.

Immunohistochemistry was performed to determine the distribution ofimmune cells within the tumor. In particular, CD11b myeloid cells wereexamined, as were regulatory vs. effector T cells (CD3, CD4, CD8,FoxP3). Briefly, IHC for CD11b or CD68+ was performed as follows. CD11bantibodies were available from Abcam, clone 1-70, Catalong # ab 8878.CD68 antibodies were available from Abcam, clone FA-11, catalog # ab53444. Tissues were fixed in cold methanol, air-dried, and then stainedusing a Res IHC Omni-UltraMap HRP XT protocol.

As a baseline, untreated murine breast cancer cells showed moderatelevels of CD11b+myeloid cells (data not shown). FIG. 37A shows that upontreatment with Compound BB, there was a decrease in CD11b+ myeloid cellstaining in viable regions of 4T1 tumors treated with Compound BBcompared to vehicle, on day 13 of study. Without being bound by aparticular theory, the decrease in the pro-tumorigenic tumor associatedmyeloid cells can result in activation of cytotoxic T-cells andanti-tumor immunity. FIG. 37B shows image analysis of tumor tissuetreated with Compound BB or vehicle, where Compound BB resulted in adecrease of CD11b+ cells in the tumor. FIG. 37C shows quantification ofthe percentage of CD11b+ cells or CD68+ cells in tumor tissue treatedwith Compound BB or vehicle. FIG. 37E and FIG. 37F show quantificationof CD11b levels by IHC on tumor cells from 4T1-luc mammary fat padtumors; Compound BB reduces the percentage of tumor cells observed. Whenthe images were quantified as CD11b+ cells per total viable cells (FIG.37E), a Student t-Test p-value of 0.0355 was obtained when comparing theCompound BB-treated cells to controls. When the images were quantifiedas CD11b+ stained area per total visible area (FIG. 37F), a Studentt-Test p-value of 0.0441 was obtained when comparing the CompoundBB-treated cells to controls. Thus, Compound BB causes a statisticallysignificant reduction in CD11b+ cells in this assay. FIG. 37G and FIG.37H show further IHC analysis showing how Compound BB affects thepercentage of CD11b-stained cells. When the images were quantified asCD11b+ cells per total viable cells (FIG. 37G), a Student t-Test p-valueof 0.0864 (d16) or 0.3967 (d22) was obtained when comparing the CompoundBB-treated cells to controls. When the images were quantified as CD11b+stained area per total visible area (FIG. 37H), a Student t-Test p-valueof 0.2074 (d16) or 0.1538 (d22) was obtained when comparing the CompoundBB-treated cells to controls. As FIG. 37D indicates, there is a strongcorrelation between CD11b+ and CD68+ myeloid cells in CompoundBB-treated 4T1 tumors.

Flow cytometry was performed to obtain a more extensive cellular subsetanalysis. In particular, the study examined T cell vs. Myeloid cells,Myeloid cell subsets, Regulatory vs. effector T cells, and Tumor markersvs. WBC (e.g., PD-L1 expression on tumor).

Different populations of tumor-associated myeloid cells can beidentified by flow cytometry, e.g., monocytic immature myeloid cells(iMC), Granulocytic iMCs/Neutrophils, Tumor Associated Macrophages(TAMs), and Granulocytic MDSCs. The different populations, and panelsfor detecting MDSCs, are described in Cancer Discovery 2011; 1:54-67.Tumors may vary in their myeloid cell composition. In LLC tumor breimodels treated with Compound BB, the number of infiltrating TAMs wasdecreased, as shown in FIG. 38A. This experiment indicates that CompoundBB successfully reduced a type of immunosuppressive cells in the tumormicroenvironment, supporting the model that Compound BB treats cancer byblocking immunosuppression at or near the tumor site. In a colon cancermodel (CT26 subcutaneous tumors) monocytic iMCs were decreased byCompound BB as shown in FIG. 38B. FIG. 39 shows that Compound BB reducedM2 polarized suppressive macrophages. CD206 is characteristic of(pro-tumor) M2 macrophages or MDSCs, and CD206 staining was reduced inLLC tumor brei samples upon treatment with Compound BB, compared to acontrol.

In conclusion, Compound BB shows a pharmacodynamic effect on myeloidcells that is consistent with an anti-tumor activity. This effect wasobserved in 4T1-luc breast cancer cells, LLC-luc lung cancer cells, andCT26 colon cancer cells.

In addition RNA profiling was performed to determine the M1 vs. M2status of differentiated macrophages, as well as broaderimmunosuppressive vs. inflammatory panel in whole tumors.

M2 macrophages can be produced as follows. Sacrifice mouse and collectfemur and tibias. Immerse the bones on a petri dish in 70% ethanol for1-2 min, then remove the bones and let the ethanol dry off for 3-4 min.Cut off a very small portion of the end of the bones to make an easyaccess for needle insert. Using a 27 g needle filled will mediacontaining DMEM+10% FBS+1% Pen/Strep (complete DMEM), expel the bonemarrow from both ends of the bones into a 15 ml petri dish. Pipette thebone marrow to dissociate the bone marrow cells. Centrifuge and add 1×Red Cell lysis buffer to the pellet. Let RBC lyse for 2 min then addequal amount of complete DMEM and centrifuge. The bone marrow from onemouse can be divided into 2 petri dishes containing 7 ml of BMM media(DMEM+20% FBS+1% Pen/Strep+50 ng/ml M-CSF). The day that the cells areplated onto petri dishes is day One. On day four add 3 ml BMM media toeach plate. Add the M-CSF right before culturing. The cells are readyfor polarization on day 6.

Next, the cells can be polarized. To harvest cells, remove thesupernatant and wash once with 1×PBS. Add 4 ml Accutase detachmentsolution per petri dish. Leave for 5-10 min and collect cells and washthe petri dish with complete DMEM to obtain all the cells on the dish. Alonger harvest time can increase yield. At this point cells can beplated for experiments.

1. Add DMEM+10% FBS+1% Pen/Strep to the cell pellets.

2. Plate cells into 12 well plates (1×10{circumflex over ( )}6 cells/2ml media per well) in complete DMEM containing 20 ng/ml M-CSF.

3. For M1 polarization add: 20 ng/ml IFNg+100 ng/ml LPS for 24 hrs

4. For M2 polarization add: 20 ng/ml IL-4 ng/ml for 48 hrs

5. Add the treatments/inhibitors at the same time.

6. Add test drug (for example test concentrations of Compound BB and+/−DMSO controls)

7. Harvest RNA.

RNA quantity and quality can be evaluated to normalize prior to RT PCR.qRT PCR is performed according to the manufactures recommendations usingthe RNA to CT one step kit from ABI and ABI taqman primer kits forrelevant genes. If the cells are being analyzed by FACS, thedifferentiated cells are lysed/fixed (bd) after harvest, and frozen.Staining for mouse CD206 (Biolegend 161707) and ARG 1 (R and D systemsIC5868F) is done in BD stain buffer with BSA.

RNA expression profiling was performed for immuno-phenotyping with aTaqman low density array (TLDA). Table 24 below shows a panel of genesthat can be used to determine the phenotype of immunoinfiltrates.

TABLE 24 Genes that can be used to determine the phenotype ofimmunoinfiltrates Tumor Cell Inflammatory T-cell Immuno-microenvironment& population tumor killing activation suppressiveangiogenesis CD45 TNFa IL6R FOXP3 PDGF CD11c IFNγ CD62L CTLA4 VEGF CD11bIL12b CD25 INOS VEGFR1 CD3 Granzyme B CD80 IDO VEGFR2 CD4 CD86 TGFbSMA1a CD8 CD83 IL10r CD68 CD137L PD-1 GITRL ARG1 CD40 PDL1 OX40L BACH2OX40 ADM

KPC Pancreatic Cancer cells were implanted into syngeneic animals whichwere subsequently treated with vehicle (n=13), Compound BB at 3 mg/kg(n=7), anti-PDL1 (n=9), or a combination of Compound BB and anti-PDL-1(n=8). For mouse tumors, snap frozen samples were powdered with aGenogrinder at liquid nitrogen temperatures and RNA was isolated withTrizol. Next, the sample was cleaned-up using Qiashredder, and thenusing a RNeasy kit.

Data was analyzed as follows. CT values were used for analysis. For eachsample Si and each gene, the following values were calculated:dCT(Si,gene)=CT(Si,gene)−CT(Si,ACTB)ddCT(Si,gene)=dCT(Si,gene)−median(dCT(vehicle treated samples,gene))RQ(Si,gene)=2{circumflex over ( )}(−ddCT(Si,gene))

This produced a matrix of RQ values: 48 genes (rows)×39 samples (cols).

Four rows were added to the RQ matrix to represent the treatment withvehicle, Compound BB, anti-PDL1, or a combination of Compound BB andanti-PDL-1. The analysis used R package Non-negative MatrixFactorization (NMF) with rank=3 and the “lee” algorithm. FIG. 40 showsnormalization to β-actin gives stable expression of GAPDH and 18S genes.

The analysis shows that M2 macrophage markers VEGF and ARG-1 trendedtowards reduction by Compound BB with or without PDL-1 antibody. Thisresult is consistent with Compound BB inhibiting the differentiation ormigration of M2 macrophages. The analysis indicates that Compound BBalone resulted in a trend in lower T regulatory and T effector genes,and a decrease in Dentritic cell activation markers. A PDL1 antibody incombination with Compound BB did not exhibit these effects. It alsoindicates that Compound BB alone or in combination with PDL1 antibodyyielded a decrease in the M2 genes ARG-1, VEGF, i.e., Compound BB seemsto reduce the level of M2 cells in the sample, for example, by blockingdifferentiation of bone marrow cells into M2 cells. As a control, PDL-1antibody resulted in the expected induction of pro-inflammatory genes.FIG. 41 shows the effect of Compound BB on markers Cd8b1, Pdcd1/PD1,Cd4, Cd3e, Foxp3, and Clla4 with or without PDL-1 antibody. FIG. 42Ashows M2 macrophage markers VEGF and ARG-1 are reduced by Compound BBwith or without PDL-1 antibody. FIG. 42B shows the effect of Compound BBwith or without PDL-1 antibody on M2 macrophage markers ARG1, VEGFa andADM.

Example 268: Physical and Pharmacokinetic Properties of Compound BB

The phyiscochemical properties of Compound BB are drug-like, andsuitable for use of the compound as a drug. For instance, Compound BBhas a molecular weight of 528.6 Da, a c Log P of 4.0, a stability in 38°simulated gastric fluid of >4 hours, and a pKa that shows no ionizationfrom pH 3-10.

ADME characteristics of Compound BB were examined, and are shown belowin Table 25.

TABLE 25 ADME characteristics of Compound BB Papp [10-6 cm/s] @ 10 uMHigh PGP efflux@ 10 uM No PGP Inhibition @ 10 uM 98% (IC₅₀ < 1 uM) PPB %free @ 1 uM 1.0-5% (all species) M, C, D, H Mic (t_(1/2) min) >120 all(incl rat) M, C, D, H Hep (t_(1/2), min) >240 all (incl rat) GSH AdductFormation Neg Human hep viability IC₅₀ 10-25 uM Rev Cyp Inhibition IC₅₀All Cyps > 10 uM Time dep Cyp Inhibition @ 10 uM Neg Cyp Induction @ 10uM Neg (activity and mRNA) Mouse PK AUC_(last) @10 mg/kg in 83742 NMPsolution (ng*h/mL)

Compound BB exhibits high cellular membrane permeability. In addition,the compound does not substantially inhibit CYP. Compound BB achieveshigh oral bioavailability in rodent and non-rodent animal species.Compound BB achieves a high volume of distribution and appears toreadily distribute into cells/tissues. It also has high metabolicstability. This stability translates to low clearance and a longhalf-life in vivo. These properties indicate the compound's suitabilityfor in vivo administration.

(Compound BB appears to be an inhibitor of P-GP (P-glycoprotein 1). Insome embodiments, a PI3K-γ inhibitor such a Compound BB is administeredto a patient in combination with a second therapeutic that is a P-gpsubstrate. Without being bound by a particular theory, the P-gpinhibitor could help maintain levels of the P-gp substrate in tumorsoverexpressing the P-gp transporter.

(The oral half-life of Compound BB is about 3.3 hours in rat, about 7.1hours in dog, and about 10.2 hours in monkey. Based on this data, theoral half-life in humans is expected to be 10-13 hours.

(There is low potential for reversible inhibition of main CytochromeP450 isoforms. Compound BB is not expected to form reactive metabolites.

In one exemplary study, the plasma concentrations of Compound BB within24 hours following oral administration in rat (7.5 mg/kg), dog (5mg/kg), and monkey (5 mg/kg) were measured and the results are shown inFIG. 43. Compound BB's oral exposure is high in rat, dog and monkey.Good oral bioavailability and low clearance in rodent and non-rodentanimal species results in high oral exposure.

In another exemplary study, the plasma exposures of Compound BB in ratsfollowing oral administration at 10, 25, and 100 mg/kg were measured (byAUC 0-last) and the results are shown in FIG. 44. The increase inexposure of Compound BB is greater than or equal to the increase indose. Thus, the compound exhibits good dose-exposure proportionality.

Example 269: Toxicology Studies of Compound BB

In vitro toxicology studies of Compound BB showed that the compound isexpected to have very low toxicity. A target organ toxicity (TOT) studywas performed in rats. The rats were dosed orally with Compound BB for10 days. This study assessed clinical observations/mortality, bodyweights, clinical pathology, toxicokinetics, and histopathology ofselect tissues. The tissues studied were: Adrenal gland, Bone marrow(femur), Brain, Small Intestine (including GALT), Heart, LargeIntestine, Kidney, Stomach, Liver, Pancreas, Spleen, Testis, and Thymus.Doses were selected based on repeat-dose tolerability study, PK andplasma protein binding, and cellular IC₅₀s for PI3K isoforms. The dosesselected were: high-dose: 50 mg/kg/day, expected to inhibit α, β, δ, andγ PI3K isoforms; Mid-high dose: 20 mg/kg/day, expected to inhibit (3, 6,and γ PI3K isoforms; Mid-low dose: 7.5 mg/kg, expected to inhibit γ PI3Kisoform only; and Low-dose: 1 mg/kg/day, expected to inhibit γ PI3Kisoform only. The formulation used was Formulation: 0.5% CMC, 0.05%Tween in water with Compound BB.

The TOT study indicated that Compound BB was well-tolerated for 10 days.No mortality or clinical signs were observed at any dose level. Therewas a minimal decrease in body weights of high dose animals (5-8%)compared to vehicle. As to clinical pathology, minimal changes inneutrophils, lymphocytes, and fibrinogen, (primarily in 20 and 50mg/kg/day groups) were observed. In terms of histopathology, nosignificant gross changes were observed. In the spleen, there wasminimal/mild lymphoid depletion of the marginal zone and periarteriolarlymphoid sheaths (PALS) primarily at doses≥20 mg/kg/day, and in thethymus, there was minimal/mild lymphoid depletion in males treated at≥7.5 mg/kg/day and in females at 50 mg/kg/day.

FIG. 45 shows the concentrations of unbound Compound BB, in a TOT study.Free Compound BB reaches concentrations that are expected to inhibitPI3K isoforms. Evaluation of the PI3K isoform inhibition data relativeto free plasma concentrations demonstrates that the doses that wereadministered inhibited the desired isoforms. Specifically, the 1 and 7.5mg/kg/doses achieved high enough concentrations to selectively inhibitPI3K γ without substantially inhibiting PI3K α, β, or δ.

Example 270: Effect of the Tumor Microenvironment on Cancer CellSurvival

Bone marrow stromal cells protect CLL cells from spontaneous apoptosisin vitro. This is shown in an experiment where CLL cells were added toconfluent stroma, and then CLL viability was assessed over time. FIG. 46shows that CLL cells have a higher survival rate when co-cultured withstromal cells. This experiment suggests that stromal cells sendpro-survival signals to CLL cells, and these signals are mediated byPI3K. Experiments with selective inhibitors for PI3K-δ and PI3K-γindicates that TME-induced CLL proliferation is dependent on PI3K-δ(data not shown).

This signaling also seems to occur in vivo, as there is an increase inPI3K signaling in proliferation centers of human CLL lymph nodes, asmeasured by high levels of pS6. Balakrishnan et al., ASH 2013. Inaddition to pS6, which is an indicator of AKT activation, theseproliferation centers have high levels of T-cells, as indicated by CD3staining (data not shown). High levels of proliferation are indicated bythe presence of Ki-67.

Example 271: Developing a Panel of Genes for Identifying MDSCs

The goal of this study is to develop a myeloid cell panel fordissociated tumor cells to measure infiltrates for TAMs and MDSCs. 4T1tumors were assayed by FACS for the presence of various immune cells.The study quantified the level of Ly6C and Ly6G in the population. Thestudy indicated that 4T1 cells are much more enriched for granulocytic(Ly6C+, Ly6G+) than LLC tumor brei models (data not shown). FACSanalysis also showed that In vitro differentiated M2 (or M1) cells havean “TAM” MDSC phenotype (data not shown).

Potential markers for a myeloid cell panel for M1 cells includes: M1CD68 MHCII, CD68 iNOS (PLoS One, 2013; 8(12): e79769).

Potential markers for a myeloid cell panel for M2 cells includes:CD68+CD163+ or CD68+ VEGF+(PLoS One, 2013; 8(12): e79769).

Potential markers for a myeloid cell panel for myocytes includes CD14and CD16, with attention paid to the ratio between the two, e.g.,CD14+CD16 —is the classical pattern, while CD low CD16 bright is nonclassical.

CSF1R is another potential marker (Cancer Cell 25, 1-14, Jun. 16, 2014^(a)2014 Elsevier Inc.). The Cancer Cell publication describes abiomarker trial in solid tumors.

Other potential markers for a panel include CD11b F4/80; CD11b LY6C/G;ARG1, CD206, and MHC expression, which can differentiate between M1 andM2 cells in mouse); and VEGF (which is highly expressed in a subset ofTAMs).

Example 272: Efficacy of Compound BB in a Glioblastoma Model

The purpose of this study is to evaluate the anti-tumor activity andimmune modulatory effects of Compound BB in the GL261-Luc murineglioblastoma subcutaneous model. GL261-luc was a murine luciferaseexpressing brain derived cell line which was stably transfected withfirefly luciferase gene. GL261-Luc adherent cells were grown in tissueculture, in filtered DMEM medium supplemented with 10% fetal bovineserum and 1% pen/step. Tumor innoculum was prepared at a concentrationof 1×107 cells in 100 uL of sterile PBS+matrigel. Cells were implantedsubcutaneously in the right hind flank area to C57BL/6 Albino male mice(stock #000058-age 6 weeks) from Jackson Laboratories. Treatment beginswhen tumors reach ˜100-200 mm³ in volume. Compound BB or vehicle wasorally administered once daily at 15 mg/kg in the NMP formulation.Efficacy was determined by tumor volume and luciferin measurements.Tumor measurements and body weights were taken two to three times aweek. Luciferin measurements were taken twice a week. At the end of thestudy, tumor cells were harvested and prepared for analysis by FACS,IHC, RNA analysis, or other analysis.

FIG. 47A shows that Compound BB reduced tumor volume in the GL-261syngeneic glioma model at 15 mg/kg QD, PO. FIG. 47B shows that CompoundBB reduced the total flux in the same model. FIG. 47C is a repeat studyof the same model that shows the effects of Compound BB QD on tumorvolume from day 9 to day 25 after implant.

Example 273: Efficacy of Compound BB with or without Cyclophosphamide inan LLC Model

The purpose of this study is to evaluate the combination effect ofCompound BB+/−cyclophosphamide in the LLC-tumor brei model. In thisexperiment, treatment began when tumors reached approx. 150 mm³—aroundday 12. Caliper and xenogen images were monitored. Treatment groupsn=13-15/group. The following doses and formulations were used: Vehicle(5% NMP/95% PEG 400/PBS) PO/IP; Compound BB @ 15 mg/kg QD, PO;Cyclophosphamide @ 50 mg/kg EOD IP; Combination (Compound BB+Cyclo).Samples for FACs analysis were collected on day 23 post implant. UsingFACS, cells were assayed for the level of CD11b and Gr-1. Gr1+CD11b+myeloid cells are the most prevalent inflammatory cells found in tumors,where they directly promote tumor angiogenesis and immunosuppression.Hardamon et. al., Proceedings: AACR 103rd Annual Meeting 2012, CancerResearch: Apr. 15, 2012; Volume 72, Issue 8, Supplement 1.

FIG. 48 shows the results of this experiment. Compound BB alone andcyclophosphamide alone each reduced tumor growth by about the sameamount. When both compounds were administered together, tumor growth wasfurther slowed. FIGS. 49A, 49B, 49C, 49D, and 49E show CD11b/Gr-1 plotswhen Compound BB was administered at 15 mg/kg and Cyclophosphamide at 50mg/kg. FIG. 49A shows the percentage of cells that are CD3 relative toCD45+. FIG. 49B shows the percentage of cells that are CD11b+Gr-1−relative to CD45+. FIG. 49C shows the percentage of cells that areCD11b+Gr-1+ relative to CD45+. FIG. 49D shows the percentage of cellsthat are CD11b-Gr-1− relative to CD45+. FIG. 49E shows the percentage ofcells that are CD11b-Gr-1+ relative to CD45+.

FIGS. 50A, 50B, 50C, and 50D show MDSC panel CD11b/Ly6C/Ly6G whenCompound BB was administered at 15 mg/kg and Cyclophosphamide at 50mg/kg. FIG. 50A shows the percentage of cells that are CD11b+ relativeto CD45. FIG. 50B shows the percentage of cells that are CD11b+Ly6C+relative to CD45+. FIG. 50C shows the percentage of cells that areCD11b+Ly6C+Ly6G+ relative to CD45+. FIG. 50D shows the percentage ofcells that are CD11b+Ly6C-Ly6G− relative to CD45+.

FIGS. 51A, 51B, 51C, 51D, and 51E show CD3/CD4/CD8 plots when CompoundBB was administered at 15 mg/kg and Cyclophosphamide at 50 mg/kg. FIG.51A shows the percentage of cells that are CD3+ relative to CD45+. FIG.51B shows the percentage of cells that are CD4+CD8− relative to CD45+.FIG. 51C shows the percentage of cells that are CD4+CD8+ relative toCD45+. FIG. 51D shows the percentage of cells that are CD4-CD8+ relativeto CD45+. FIG. 51E shows the percentage of cells that are CD4-CD8−relative to CD45+.

Example 274: The Effect of Compound BB on CD206 Levels of Macrophages inLLC Tumors

The purpose of this study is to evaluate the effects of Compound BBdosed PO, QD along or in combination with cyclophosphamide. C57 Albinomice were dosed QD with Compound BB. Cyclophosphamide was dosed EOD IP.Tumor measurements and bodyweights were taken 2×/week. Tumors wereharvested on the last day or if the vehicles reached approx. 3000 mm³.Tumors were cut in half. One half was cut in half again with ½ fixed in10% NBF, and the other was frozen in OCT for frozen sections. Theremaining half was processed to single cell suspension and evaluated byFACS using two panels.

Antibodies used for staining for FACS were as follows.

Staining with CD4 and CD8a: CD45-AF700 (BDB560510) at 1.25 ug/ml;CD3-FITC (BDB555274) at 1 ug/ml; CD4-PE (BDB553049) at 2.5 ug/ml; andCD8a-PerCP (BDB553036) at 5 ug/ml.

Staining with CD11b and GR-1: CD45-AF700 (BDB560510) at 1.25 ug/ml;CD3-FITC (BDB555274) at 1 ug/ml; CD11b-APC (BDB553312) at 2.5 ug/ml; andGR-1-PE-CY7 (BDB552985) at 0.5 ug/ml.

Staining with CD206 panel: CD45-AF700 (BDB560510) at 1.25 ug/ml; CD11bPE at 2.5 ug/ml; and CD206 APC at 25 ug/ml.

Staining with MDSC panel: CD45-AF700 (BDB560510) at 1.25 ug/ml; CD11b PEat 2.5 ug/ml; Ly6C APC at 2.5 ug/ml; and Ly6G FITC at 2.5 ug/ml.

FIGS. 52A, 52B, 52C, 52D, and 52E show the percentage of cells that areCD45+, CD11b+, or CD206+, expressed as a percent of total cells, in LLCtumor brei model treated with Compound BB+/−cyclophosphamide. FIG. 52Ashows the percentage of cells that are CD45+ expressed as a percent oftotal cells. FIG. 52B shows the percentage of cells that are CD11bexpressed as a percent of total cells. FIG. 52C shows the percentage ofcells that are CD206 expressed as a percent of total cells. FIG. 52Dshows the percentage of cells that are CD11b expressed as a percent oftotal cells. FIG. 52E shows the percentage of cells that are CD206expressed as a percent of total cells.

There appears to be a decrease in the CD206 population with bothCompound BB administered alone and in combination with cyclophosphamide.Compound BB therefore reduces the number of M2 macrophages in the tumor,either by preventing them from migrating into the tumor or by preventingimmature myeloid cells from differentiating into M2 cells, or both.

Example 275: Pharmacokinetics and Reduction of LPS-InducedNeutrophilia—Intratracheal Administration

The study of intratracheal (it) administration of Compound BB onLPS-induced neutrophilia model is carried out using methods known in theart. For example, Compound BB is administered intratracheally to rats atan amount of e.g., 1 μg/kg, 10 μg/kg or 100 μg/kg. The administration ofthe controls is performed as e.g.: (1) oral administration of CompoundBB at 10 mg/kg; (2) intratracheal administration of saline (vehicle) at0.25 mL/rat; (3) intratracheal administration of LSP (vehicle) at 1 μgLPS/rat; and (4) intratracheal administration of budenoside at 0.3mg/kg. At one hour post-administration, neutrophilia is induced bytreatment with LPS, and the samples are collected by bronchoalveolarlavage (BAL) at varying hours post-administration and subjected tofurther examination. Compound BB is a PI3Kγ inhibitor that has adelta/gamma selectivity ratio of greater than about 50. As a comparison,similar procedures are carried out using a PI3K-δ inhibitor, which has agamma/delta selectivity ratio of greater than about 50.

The levels of Compound BB in lung, bronchoalveolar lavage fluid (BALF),and plasma are determined using methods known in the art.

Example 276: Studies with Smoking Mouse Model of Chronic ObstructivePulmonary Disease (COPD)

Studies of the effect of the compounds provided herein on smoking mousemodel of COPD can be conducted using methods known in the art.

A. Provided below is an illustrative example of such method. The miceare exposed to cigarette smoke for 4 days in a whole body exposure box.Whole body exposure is conducted for a certain amount of time in acustom made cylindrical 32 L Perspex box (e.g., Boehringer IngelheimPharma GmbH & Co. KG, Biberach, Germany) before each cigarette smokeexposure or before lung function measurement. Animals are separated bystainless steel spacers. The floor of the box is additionally heated(e.g., 38° C.) to maintain the physiological body temperature of theanimals. Control animals receive solvent as placebo. On day one and two,mice are exposed to the mainstream smoke of cigarettes. Exposure to thesmoke of each cigarette lasts for about 15 min followed by an 8 minexposure with fresh room air. Every second cigarette an additional breakof 24 min with exposure to fresh room air is conducted. A semi-automaticcigarette lighter and smoke generator with an electronic timer is usedto control the cigarette exposure (e.g., Boehringer Ingelheim PharmaGmbH&Co. KG, Biberach, Germany). Cigarette smoke particle concentrationis monitored by a real time ambient particle monitor (e.g.,MicroDustPro, Casella, Amherst, N.H., USA). Control animals are exposedto room air. See e.g., L. Wollin, et al., Pulmonary Pharmacology &Therapeutics 23 (2010) 345-354.

Compounds provided herein are administered orally or intra-tracheally(it) using, e.g., methods know in the art. For example, a compoundprovided herein is administered orally as a solid or a solution.Alternatively, a compound provided herein is administeredintra-tracheally by aerosolizing a solution of the compound with a jetnebulizer. A compound provided herein is administered at variousconcentration and schedules. For example, whole body exposure of acompound provided herein is administered for 5 min and 1 h prior toexposure to cigarette smoke.

Differential cell counts are determined in the bronchoalveolar lavagefluid (BALF). For example, the total BAL cell counts and the amount ofneutrophils in the samples are used to evaluate the efficacy of acompound provided herein. Models of cigarette-smoke induced pulmonaryinflammation have an increased BAL total cell count and also the amountof neutrophils. As such, a decrease of BAL total cell count and theamount of neutrophil in models administered with a compound providedherein as compared to a control model (without administration of acompound provided herein) illustrates the effectiveness of a compoundprovided herein in the treatment of pulmonary inflammation. See e.g., L.Wollin, et al., Pulmonary Pharmacology & Therapeutics 23 (2010) 345-354.

Further, lung sections stained with e.g., H&E or AB/PAS of the controlmodels (e.g., models without administration of a compound providedherein) are compared with lung sections of models administered with acompound provided herein. H&E staining shows inflammation and alveolarinfiltrates in the lungs. AB/PAS staining shows the mucus content in thegoblet cells of the large airway. See e.g., L. Wollin, et al., PulmonaryPharmacology & Therapeutics 23 (2010) 345-354.

B. Provided below is an example for a COPD 10 days cigarette smokemodel. The goal of this study is to determine whether treatment with acompound provided herein can prevent the steroid resistant pulmonaryinflammation induced by exposing mice for 10 days to cigarette smoke.Mice are first exposed for 5 days to cigarette smoke to induceglucocorticosteroid resistance. Thereafter, the animals are exposed foran additional 5 days to cigarette smoke together with the administrationof the test compounds to investigate whether the glucocorticosteroidresistance can be diminished.

Specifically, Balb/c byJ mice (weight 24-26 gr; 10-12 weeks old) areexposed to standard air or cigarette smoke for 10 days (“whole bodyexposure”). Mice are exposed when they are sitting together in onePerspex box. The cigarettes used are special research cigarettesstandardized from Kentucky University: 3R4F without filter. Dose andtime are increased starting at the 1^(st) till the 10^(th) day ofexposure (twice a day (=2 runs); interval exposure is 5 hours):

-   -   1st day: run 1=2 pairs of cigarettes and run 2=3 pairs of        cigarettes. The smoke exposure dose and time=10-15 min. (CO        dose=150-300 ppm; O₂ concentration=20.8% measured by Baccharach        PCA3-analyzer);    -   2nd day: run 1=4 pairs of cigarettes and run 2=5 pairs of        cigarettes. The smoke exposure dose and time=20-25 min. (CO        dose=150-300 ppm; O₂ concentration=20.8%    -   3rd day: run 1=6 pairs of cigarettes and run 2=7 pairs of        cigarettes. The smoke exposure dose and time=0-35 min. (CO        dose=150-300 ppm; O₂ concentration=20.8%;    -   4th day till the 10th day: run 1=7 pairs of cigarettes and run        2=7 pairs of cigarettes. The smoke exposure dose and time=35        min. (CO dose=150-300 ppm; O₂ concentration=20.8%.

Animals are treated with a compound provided or Vehicle solution 1 or 2orally daily from day 6 till 10. Dexamethasone (5 mg/kg) is administereddaily intraperitoneally (IP) from day 6 till 10. Body weight of theindividual animals is monitored daily from day 1 till day 11.

Mice are sacrificed 1 day after the last exposure to air or CS (day 10).Blood is collected via a heart puncture and the isolated serum iscollected and stored at −30° C. Lungs are lavaged, BAL cells areisolated counted and differentiated. BAL fluid is stored fordetermination of cytokines/chemokines.

Below are tables of the experimental setup:

Day Treatment 1 till 10 Balb/c byJ mice (weight 24-26 gr; 10-12 weeksold) are exposed to standard air or cigarette smoke. 6 till 10 Animalsare treated with a compound provided herein or Vehicle solution 1 or 2orally daily from day 6 till 10. Dexamethasone (5 mg/kg) is administereddaily intraperitoneally (IP) from day 6 till 10. 1 till 11 Body weightof each individual mouse is noted once a day. 11 The animals aresacrificed. Blood is isolated. Serum is stored for cytokine/chemokineprofiling (optional). Lungs are lavaged, BAL cells are isolated,counted, and differentiated. BAL fluid is stored for determination ofcytokines/chemokines (optional).

Treat Veh TC group Model Veh 1 2 Dex 1 TC2 TC3 N = 1 Air X X 10 2Cigarette Smoke X X 10 3 Cigarette Smoke X X 10 4 Cigarette Smoke X X 105 Cigarette Smoke X X 10 6 Cigarette Smoke X X 10 7 Cigarette Smoke X X10 8 Cigarette Smoke X X 10 9 Cigarette Smoke X X 10 Total 90

C. Provided below is a murine smoking model. See e.g., Yasuo, T., etal., Am J Respir Crit Care Med 2010; 182:897-904; Sato T, et al., Am JRespir Crit Care Med 2006; 174:530-537; and Kasagi S, et al., Am JPhysiol Lung Cell Mol Physiol 2006; 290: 396-404.

Inhalation of tobacco smoke is performed utilizing unfiltered researchcigarette 1R1 (Tobacco Health Research Institute, Kentucky Univ.,Lexington, Ky.) and the Tobacco Smoke Inhalation Experiment System forSmall Animals (model SIS-CS; Shibata Scientific Technology, Tokyo,Japan). Model SIS-CS consisted of both a tobacco smoke generator (modelSG-200) and an inhalation chamber. The smoke generator is controlled bya laptop computer and automatically generated tobacco smoke by setting avolume of syringe pump (10-50 cm³/puff) and a number of puffs/min (1-12puffs).

The tobacco smoke generated is delivered to the inhalation chamber, towhich the mouse body holders are set (maximum 12 body holders can be setat a time), and mice inhale tobacco smoke through their noses. Tobaccosmoke can be diluted to a desired concentration at the blender, wheretobacco smoke and compressed air are combined, before delivery to theinhalation chamber. At 12 wk of age, mice are subjected to theexperiment of chronic inhalation of tobacco smoke. The experimentalsettings are as follows: 15 ml of stroke volume and 12 puffs/min togenerate tobacco smoke, 1.5% tobacco smoke diluted by compressed air.The mass concentration of total particulate matter (TPM) in 1.5% tobaccosmoke is determined by gravimetric analysis of filter samples takenduring exposure periods and is 23.9 mg of TPM/m³.

First, mice are trained to be set into a body holder for 30 min/daywithout smoking for 5 days, followed by exposure to 1.5% tobacco smokefor 15 min/day for 5 days. After this induction period, mice continue toinhale 1.5% tobacco for 30 min/day, 5 days/wk, and for 8 wk. As acontrol for the inhalation of tobacco smoke, mice are subjected to thesame experimental procedure with the same conditions described above,but air was delivered instead of tobacco smoke.

Example 277: Lung Inflammation Assay

Compounds provided herein can be tested using one or both of theLPS-induced lung inflammation assay and the ovalbumin-induced lunginflammation assay.

To perform the LPS-induced lung inflammation assay, compounds are dosedorally. A group is dosed with vehicle only and dexamethasone is used inanother group as positive control. Pulmonary inflammation is determined6 h after intranasal instillation of LPS. The following parameters canbe evaluated: total number of leukocytes and number of neutrophils inbronchoalveolar lavage (BAL).

In the ovalbumin-induced lung inflammation assay, compounds are dosedorally. A group is dosed with vehicle only and dexamethasone is used inanother group as positive control. Pulmonary inflammation is determined4 days after 4 consecutive daily intranasal instillation of ovalbumin.Compounds are given by gavage 30 min before each challenge (4challenges) at the indicated doses. The following parameters can beevaluated: total number of leukocytes and number of eosinophils inbronchoalveolar lavage (BAL).

Example 278: Efficacy of Compound BB with or without Temozolomide inGlioma Model

The purpose of this model is to evaluate the GL-261 glioma model treatedcontinuously with Compound BB or with temoloamide (TMZ) followed byvehicle or Compound BB. TMZ was dosed IP 1×/day for 7 days. The groupthen were randomized to receive Compound BB @ 15 mg/kg QD dosed PO orvehicle to evaluate the delay in tumor regrowth.

Animals: C57 Albino mice (male) from Jackson Labs, n=80. ParametersEvaluated:

In life: Mice were dosed QD with Compound BB. Temozolomide was dosed QDIP, for 7 days. Tumor measurements and bodyweights were taken 2×/week,but caliper and xenogen measurements.

End point analysis: Tumors were cut in half. One half was cut in halfagain with ½ fixed in 10% NBF the other was frozen in OCT for frozensections. The remaining half was processed to single cell suspention andevaluated by FACS.

PD satellite group—8 mice from the TMZ treated mice and 8 mice from thevehicle treated mice were sacrificed on day 7 post treatment. Tumorswere processed to single cell suspention and evaluated by FACS.

The study overview is shown in the following Table 26:

TABLE 26 Dose Dose Drug Grp (mg/ vol conc # N Compound kg) RouteFrequency (ml/kg) (mg/ml) 1 12 Vehicle 0 PO/IP QD 5/10 0 2 12 CompoundBB 15 PO QD  5 3 3 20 Compound BB 15 PO QD  5 3 4 40 Temozolomide 1 IP10 5 4-PD 8 Vehicle 0 PO/IP QD 5/10 0 4-PD 8 Temozolomide 1 IP QD 5/10 0

FIGS. 53A, 53B, 53C, 53D, and 53E show the effect of Compound BBfollowing temozolomide treatment on GL-261 tumors. FIG. 54A shows theeffect of TMZ treatment versus vehicle and FIG. 54B shows the effects ofCompound BB on tumor volume post TMZ treatment. Compound BB is effectivein reducing the tumor volume in GL-261 tumors following the treatment oftemozolomide.

Treatment with TMZ @ 1 mg/kg followed by treatment with Compound BBresulted in significant reduction in tumor volume as compared to vehicletreated mice. Mice treated with Compound BB from the start of treat alsoresulted in a significant reduction in tumor volume as compared tovehicle treated mice. The two groups were not significant from oneanother but with longer treatment and monitoring this may result in asignificantly bigger window.

Example 279: Effect of Compound BB and Anti-PDL-1 on T CellProliferation with CT26 Tumors

The goal of this study is to evaluate the effect of Compound BB on Tcell proliferation with CT26 tumors. CT26 was anN-nitroso-N-methylurethane-(NNMU)induced, undifferentiated coloncarcinoma cell line. It was cloned to generate the cell line designatedCT26.WT (ATCC CRL-2638).

The duration of the study was 1 week under treatment. 5-6 week oldfemale, Balb/c mice (Jackson Labs) implanted subcutaneously with5×10⁵CT26 cells in 100 μl PBS. CT26 cells were cultured in RPMIsupplemented with 10% FBS. Dosing commenced when tumors reached anaverage volume of 500-600 mm³.

The groups were treated as follows:

Vehicle Compound BB αPD-L1 PO PO IP QD QD X1 Group # mice/group (n)mg/kg mg/kg mg/kg 1 10 0 0 0 2 10 0 15 0 3 10 0 0 RTU* Group 1: Vehicle(5% NMP/95% PEG) Group 2: Compound BB prepared at 10 mg/ml Group 3:*αPD-L1 antibody comes ready to use (RTU) (1 mg/ml) dosed with 200 ug,once. Supplied by BioLegend (Cat # 124318, lot #B179643), clone 10F.9G2

For groups 1 and 2 (vehicle and compound BB, respectively), the micewere dosed QD, PO for 5 days at a volume of 5 ml/kg. For group 3(αPD-L1), the mice were dosed once, IP, 200 ug (200 uL from 1 mg/mlstock).

Mice were monitored daily for adverse clinical effects. Body weights andtumor luminescent were measured two times per week. At the end of thestudy, tumors were harvested and cut into 3 parts; one piece for CTLkilling assay, one piece for CSFE proliferation assay and one piece(split into two) for OCT/FFPE.

Spleens from 3 vehicle treated tumor bearing mice were physicallydissociated in T cell media and passed through a 70 micron filter. Afterred cell lysis, the cells were pooled, brought up in PBS and stainedwith CFSE (invitrogen). After washing, these cells were brought up in 20ml T cell media. Half the cells received Dynal mouse activator beads(CD3/CD28 crosslinking at a 1:1 concentration). CD3/CD28 beads induceproliferation in splenocytes from tumor bearing mouse (data not shown).

Tumors from vehicle treated, Compound BB treated, and anti-PDL1 treatedmice were minced, treated with collagenase and Dnase for 30 minutes at37 degrees ° C., and cells were isolated by passing the digested tumorthough a 70 micron filter. The cells were treated with red cell lysisbuffer, washed, and re-suspended in 10 ml T cell media. 2 mls of theeach tumor cell suspension were plated into each of two 12 well plates.To one of the plates, Dynal bead activated CSFE labeled splenocytes wereadded and un-activated CSFE labeled splenocytes were added to the other.CSFE labeled splenocytes were also added to wells that had T cell mediawithout tumor cells.

After four days, the cells were harvested from the wells and stainedwith antibodies for CD3, CD4, and CD8. Analysis was by FACS, measuringthe percentage of cells that remained in an un-proliferated state (CSFEstain undiluted) after 4 days. Measurements of splenocyte proliferation,either activated or un-activated, from the wells without the tumor cellsprovided the positive and negative controls for proliferation.

FIGS. 55A, 55B, and 55C show the effect of Compound BB and anti PDL-1 ontumor cells inhibition of T cell proliferation versus vehicle. FIG. 55Ashows the effect of tumor cells from vehicle treated mice. FIG. 55Bshows the effect of tumor cells from Compound BB treated mice. FIG. 55Cshows the effect of tumor cells from anti PDL-1 treated mice. FIG. 56Aand FIG. 56B show that tumor cells from Compound BB and anti-PDL-1treated mice are less inhibitory of spleen T cell proliferation ascompared to tumor cells from vehicle treated mice.

FIG. 57 shows Compound BB reduces immune suppressive environment via anex vivo T-cell proliferation assay.

In summary, T cell proliferation can be suppressed by cells derived fromCT26 mouse tumors, but treatment with either Compound BB or anti-PDL1can release the suppression.

Example 280: Compound BB Inhibits the Ex Vivo Whole Blood Stimulation ofPhospho-AKT in T Cells and Monocytes by CXCL12

C57 BL/6 Albino mice were implanted with 1×10⁶ cells in a SC injectionin the hind flank. The cell implant material was generated from breithat was passaged from mouse to mouse. Treatment started when tumorswere between 160-180 mm³. Mice were dosed for 12 days, QD, PO withvehicle, 15, 7.5 or 3 mg/kg. On day 12 mice were dosed and terminalblood was collected 2 h post dose by cardiac stick collection. Blood wasplaced in EDTA K2 collection tube and stored at room temp for PDanalysis.

Blood from each animal was aliquotted into 2 plates and warmed to 37° C.degrees. PBS was added to one plate as an unstimulated control. MurineCXCL12/SDF-1 was added to the second plate to a final concentration of800 ng/mL and plate was returned to the 37° C. degree block. After 2minutes of incubation, blood was lysed and fixed using paraformaldehyde.Samples were washed and frozen until flow cytometric analysis. T cellswere gated using an antibody to CD3 and monocytes were gated as LY6G⁻LY6C^(high). Intracellular pAKT S473 was measured and percent positivewas used for graphing. FIG. 58 shows LLC-bearing mice treated withCompound BB inhibits the ex vivo whole blood stimulation of phosphor-AKTin T cells. FIG. 59 shows LLC-bearing mice treated with Compound BBinhibits the ex vivo whole blood stimulation of phosphor-AKT inmonocytes. FIG. 60 shows the effect of Compound BB on CXCL12- oranti-IgD-induced pAKT in B cells.

Example 281: Single Dose Dog PK of Compound BB

The PK of Compound BB in male beagle dog was measured at four differentdosage: 2 mg/kg, 5 mg/kg, 10 mg/kg, and 25 mg/kg. FIG. 61 shows thesingle dose unbound plasma concentration of Compound BB versus timeprofiles in male beagle dogs.

Example 282: Bone Marrow Derived Macrophages (BMDMs) Migration Assay

BMDMs are obtained from bone marrow isolated from femurs and tibias ofC57Bl/6J mice. Bone marrow cells are cultured in a petri plate in thepresence of RPMI additioned with 30% L-cell conditioned medium(supernatant of 5 days culture of L929 cells), 20% FBS and penicillinand streptomycin. After about 10 days, almost all cells become attachedBMDM. For the migration assay, BMDMs are pretreated with 6 differentconcentrations of a compound provided herein and migration assay isperformed by using a Boyden chamber toward C5a chemokine (50 nM) orCXCL12. Migrating BMDM, attached to the polycarbonate of the Boydenchamber, are counted at the microscope (5 fields/sample) and % ofmigrating cells is calculated by relating the number of the treatedcells with the control vehicle. All values are plotted in a doseresponse curve and the EC₅₀ is defined by using nonlinear regressionanalysis (GraphPad Prism program). Compound BB can be tested for itseffect on migration of the BMDMs to C5a or CXCL12.

Example 283: Combination with Temazolamide and Radiation Therapy in theEvaluation of Orthotopic Tumor Growth of GL-261 Cells in C57Bl/6 Mice

The purpose of this study is to evaluate the effect of compound provideherein in combination with temazolamide and radiation therapy inorthotopic tumor growth of GL-261 Cells in C57Bl/6 mice.

Sixty (60) 4-6 week old C57Bl/6 male mice are anesthetized with xylazine(5 mg/kg) and ketamine (100 mg/kg) given by intraperitoneal injectionand the surgical area of the skull prepped with alcohol and Betadine. Adiagonal incision is made from behind the eye to the posterior of thehead. The bregma is exposed and the cells implanted 2 mm to the rightand 1 mm anterior to the bregma using a small drill to generate the holefollowed by a Hamilton Syringe to administer 1×10⁵ cells in a totalvolume of 10 μL a rate of 1 μL per minute. The skin is sutured and themice is recovered on a heating pad then returned to their cages. Allanimals receive post-operative pain medication every 12 hours for 72hours.

Animals are IVIS imaged 5 days after surgery to determine the total fluxper animal for randomization. After randomization, animals in Group 1are dosed with Vehicle (route/schedule TBD) and animals in Group 2 aredosed with compound provided herein, e.g., Compound BB at 15 mg/kg.Animals in Group 3 are dosed with compound provided herein, e.g.,Compound BB, and 3 fractions of radiation of 2Gy each focused to thearea of the brain tumor. Animals in Group 4 receive treatment with 25mg/kg Temazolamide (i.p.) 5 times per week on a schedule of Day 0, 1, 3,5 and 6 and are given three (3) 2Gy fractions of focused radiation tothe tumor on Days 0, 2 and 4 in addition to a compound provided hereine.g., Compound BB at 15 mg/kg. Animals in Groups 5 and 6 receive 25mg/kg Temazolamide (i.p.) 5 times per week on a schedule of Day 0, 1, 3,5 and 6 and are given three (3) 2Gy or 4Gy fractions of focusedradiation to the tumor on Days 0, 2 and 4.

For focused radiation, a lead shield containing a thin window cut out ofthe top is placed over the animal exposing only the top of the head inthe area of the tumor. On days where mice are receiving radiation, theyare anesthetized using xylazine (5 mg/kg) and ketamine (100 mg/kg) givenby intraperitoneal injection. The mice are placed on a 4-mm polymethylmethacrylate plate. Radiation is generated by a 160 kVp (15-ma) X-raysource at a focal distance of 25 cm, hardened with a 0.35 mm Cufiltration system at a rate of 1 Gy/minute. Animals are weighed andmonitored for general health daily and assigned a body condition score.Tumors are measured via IVIS imaging twice weekly beginning on Day 5 andtotal flux is determined for each animal. Experimental details aresummarized below in table below:

Temazolamide Number (i.p.) of Tumor Treatment Dose/ Day FractionatedRadiation Group Animals Cells (i.p.) Frequency 0, 1, 3, 5 & 6 RadiationFrequency 1 10 GL-261 Vehicle — — None — 1 × 10⁵ 2 10 GL-261 Compound QD— — — 1 × 10⁵ provided 15 mg/kg herein 3 10 GL-261 Compound QD TMZ 2 GyDay 0, 2 1 × 10⁵ provided 15 mg/kg 25 mg/kg and 4 herein 4 10 GL-261Compound QD TMZ 4 Gy Day 0, 2 1 × 10⁵ provided 15 mg/kg 25 mg/kg and 4herein 5 10 GL-261 — — TMZ 2 Gy Day 0, 2 1 × 10⁵ 25 mg/kg and 4 6 10GL-261 — — TMZ 4 Gy Day 0, 2 1 × 10⁵ 25 mg/kg and 4

GL-261 cells are cultured at subconfluence in DMEM growth mediumcontaining 10% FBS. Cells are administered via intracranial inoculationby first generating a hole in the skull using a small drill followed byinjection of cells 3 mm into the cranium using a Hamilton Syringe at1×10⁵ cells in a ˜10 μL volume of serum free media at a rate of 1 μL perminute.

Mice are anesthetized using xylazine (5 mg/kg) and ketamine (100 mg/kg)given by intraperitoneal injection. The mice are placed on a 4-mmpolymethyl methacrylate plate. A lead shield containing a thin windowcut out of the top is placed over the animal exposing only the area ofthe cranium to be irradiated. The tumors are directly irradiated with 2or 4 Gy of radiation on Days 0, 2 and 4. Radiation will be generated bya 160 kVp (15-ma) X-ray source at a focal distance of 25 cm, hardenedwith a 0.35 mm Cu filtration system at a rate of 1.0 Gy/minute.

Tumor progression is evaluated in life twice weekly by IVIS imaging.Mice are injected via intraperitoneal injection with 0.1 ml/20 g bodyweight of 15 mg/ml D-luciferin-K⁺ bioluminescent substrate in PBS. Ten(10) minutes following injection, mice are anesthetized under isofluraneand placed in the IVIS® Lumina at maximum sensitivity for up to 5minutes exposure to detect bioluminescence with an open emission filter.Saved images are loaded into Living Image® analysis software and colorscales matched based on maximum and minimum radiance(photons/second/cm²/steradian). Identical regions of interest are drawnaround each animal and total flux is determined in terms ofphotons/second for each region of interest.

In addition to measuring tumors in the animals in-life, animals areassigned a body condition score 3 times each week. Animals with a bodycondition of lower than BC2 is sacrificed. The conditions arecategorized as follows:

-   BC5: The animal is obese, smooth and bulky. One is unable to    identify its bone structure under the flesh and fat. Often mice in    this condition cannot groom well and hair coat may appear oily and    stained.-   BC4: The animal is over-conditioned and vertebrae are only palpable    with firm pressure.-   BC3: The animal is well-conditioned. Vertebrae and pelvis are    palpable.-   BC2: The animal is under-conditioned. Segmentation of the vertebral    column is evident and pelvis is palpable.-   BC1: The animal is emaciated, skeletal structure very prominent with    little flesh cover. Vertebrae are distinctly segmented.

All animals are weighed daily throughout this study. Group weight changeis expressed as a mean percent weight change. Animal deaths in thismodel generally occur as a consequence of anesthesia overdose or drugtoxicity. Animals are monitored on a daily basis and those exhibitingweight loss greater than 30%, are unable to ambulate, achieve food andwater, and/or appear moribund are euthanized. If the tumors appear to beulcerated, the animal is euthanized. Any adverse effects orunanticipated deaths are reported to the veterinarian immediately.

Statistical differences between treatment groups are determined usingappropriate statistical techniques. A one-way ANOVA or ANOVA on ranksare used to evaluate the area-under the curve for weight gain and tumorradiance.

Example 284: Role of T Cells in Tumor Growth Suppression

The purpose is to study the role of T cells on the tumor growthsuppression after treatment with a compound provided herein (e.g.,Compound BB).

Method A: Tumors cells are implanted into RAG−/− mice. These mice aredefective in their ability to produced either T or B cells and cannotachieve immunological memory. Tumor cells are implanted into WT orRAG−/− mice on the appropriate strain background (for CT26 colon tumorcells, BALB/C). The tumor bearing animals are randomized into vehicleand those treated with a compound provided herein (e.g., Compound BB)groups for both WT and RAG−/− mice. Efficacy is evaluated by comparingtumor growth kinetics in the WT compared to RAG−/− mice.

Method B: This method uses antibodies against the T cell markers CD4 andCD8 to deplete T cell subsets. LLC-Brie, CT26 colon cancer or MC38 cellsis implanted into the appropriate host strain of mice. Mice withestablished tumors will be randomized into treatment groups as describedbelow.

CD4 and CD8 Depletion Antibodies

aCD4/CD8 Isotype Control: Rat IgG2b Isotype (BioXcell Clone: LTF-2 Cat:BE0090-R005 Lot: TBFO)

aCD4: Diluted to 1 mg/ml in Endotoxin-Free PBS (BioXcell Clone: GK1.5Cat: BE0003-1 Lot: TBFO)

aCD8: Diluted to 1 mg/ml in Endotoxin-Free PBS (BioXcell Clone: YTS169.4 Cat: BE0117 Lot: TBFO)

Dosing Regimen

αCD⁴/aCD8 therapies will be delivered i.p. every 3 days (100 ug each)

Study Design

Vehicle; Vehicle+anti CD4; Vehicle+anti CD8; Vehicle+anti CD4/CD8;

compound provided herein (e.g., Compound BB);

compound provided herein (e.g., Compound BB)+anti CD4;

compound provided herein (e.g., Compound BB)+anti CD8;

compound provided herein (e.g., Compound BB)+anti CD4/8.

Example 285: Osteolytic Bone Cancer in Bone Cancer Model

To evaluate the effect of compound provided herein (e.g., Compound BB)on osteoclast development and whether it would prevent bone damage inthe NCTC-2472 model. The NCTC-2472 tumor cell line was propagated incell culture and harvested while in logarithmic growth. 100,000 cellswere implanted directly into the medullary cavity of the distal femur onstudy day 0.

On study day 0 the mice was weighed and randomized by body weight intotreatment groups. Mice were anesthetized with isoflurane and right kneearthrotomy performed. Cells were injected in 20 ul with a 29 gageneedle. The entry hole in the bone was filled with bone wax to preventtumor cell extravasation.

On Day 2 through 22 drug treatment was done in the following groups:Vehicle, Compound BB (15 mg/kg), and Resendronate as a control. Weightwas taken twice weekly. On study Day 22 the mice was exsanguinated underisoflurane and femur samples were taken for X ray and histopathologyanalysis. The relevant endpoints include bone erosion and abnormal boneformation.

The present invention is not to be limited in scope by the specificembodiments described herein. Indeed, various modifications of theinvention in addition to those described will become apparent to thoseskilled in the art from the foregoing description and accompanyingfigures. Such modifications are intended to fall within the scope of theappended claims. Various publications, patents and patent applicationsare cited herein, the disclosures of which are incorporated by referencein their entireties.

What is claimed is:
 1. A method of treating a solid tumor in a subject,comprising administering to the subject a therapeutically effectiveamount of a compound of the formula:

or a pharmaceutically acceptable salt thereof, in combination with ananti-PD-1 antibody, wherein the solid tumor is melanoma, gall bladdercancer, triple negative breast cancer, adrenal gland cancer, ormesothelioma.
 2. The method of claim 1, wherein the anti-PD-1 antibodyis pembrolizumab.
 3. The method of claim 1, wherein the anti-PD-1antibody is nivolumab.
 4. The method of claim 1, wherein the solid tumoris melanoma.
 5. The method of claim 4, wherein the melanoma is relapsedafter or refractory to a prior therapy, and wherein the prior therapy isan immune checkpoint therapy.
 6. The method of claim 1, wherein thesolid tumor is gall bladder cancer.
 7. The method of claim 1, whereinthe solid tumor is triple negative breast cancer.
 8. The method of claim7, wherein the triple negative breast cancer is naive to immunecheckpoint therapy.
 9. The method of claim 8, wherein the immunecheckpoint therapy is a treatment by a PD-1 or PD-L1 inhibitor.
 10. Themethod of claim 1, wherein the solid tumor is adrenal gland cancer. 11.The method of claim 1, wherein the solid tumor is mesothelioma.
 12. Themethod of claim 11, wherein the mesothelioma is epithelioidmesothelioma, sarcomatoid mesothelioma, or biphasic mesothelioma. 13.The method of claim 1, wherein the compound is administered at a dose ofabout 1-60 mg once daily.
 14. The method of claim 1, wherein thecompound is administered at a dose of about 10, about 15, about 20,about 25, about 30, about 35, about 40, about 45, about 50, or about 60mg once daily.
 15. The method of claim 14, wherein the compound isadministered at a dose of about 30 mg once daily.
 16. The method ofclaim 14, wherein the compound is administered at a dose of about 40 mgonce daily.
 17. The method of claim 1, wherein the anti-PD-1 antibody isadministered at a dose of 1 to 5 mg/kg every 2 weeks.
 18. The method ofclaim 17, wherein the anti-PD-1 antibody is administered at a dose ofabout 3 mg/kg every 2 weeks.
 19. The method of claim 1, wherein thesubject has a high number of MDSC, compared to a reference value.